Pub Date : 2024-10-02DOI: 10.1093/clinchem/hvae106.380
T Oba, M Miyachi, Y Taketani, M Mizuguchi
Background ApoAST/ALT, which have lost a coenzyme such as pyridoxal phosphate (PALP), are normally present at low levels in serum. To form the holoenzyme, IFCC recommends adding PALP into AST/ALT reagents. However, commercial reagents often do not use PALP due to its instability. In recent years, pyridoxamine phosphate (PAMP), a more stable coenzyme, has been marketed. However, the differences in holo formation ability and reagent stability between their coenzymes are not well understood. In this study, we focused on the preparation of PAMP and recombinant human ApoAST/ALT (rhApoAST/ALT), followed by fundamental investigations into their use as AST/ALT reagents. Methods PAMP was synthesized from pyridoxamine through an enzymatic reaction using pyridoxal kinase. The reaction product was purified to achieve a purity of >99% in HPLC. rhApoAST/ALT were expressed in Escherichia coli and purified with chromatography. AST/ALT assay was performed on a Hitachi 7180 automatic analyzer. PALP reagents for AST/ALT were designed to correspond with the final concentration of the IFCC method. PAMP reagents were prepared with PAMP instead of PALP. To evaluate the holo formation ability of PALP and PAMP reagents, we measured a dilution series of rhApoAST/ALT. To assess stability, the reagents were stored at 11°C for 3 months and then measured same. The reagents were subjected to content analysis by HPLC. Results Similar results were shown between coenzymes in the evaluation of holo formation ability, and high correlations were observed. As for the stability test, the values of PALP reagents decreased after 3 months, while the values of PAMP reagents did not. Content analysis showed that a spontaneous transamination occurred in PALP reagents. These results were observed at both AST and ALT reagents. Conclusions In AST/ALT reagents, both PAMP and PALP can activate apoenzymes. While PAMP is stable, PALP causes a decrease in reagent performance due to a spontaneous transamination.
{"title":"B-016 Evaluation of pyridoxamine phosphate (PAMP) for AST/ALT reagents with recombinant human apoenzymes","authors":"T Oba, M Miyachi, Y Taketani, M Mizuguchi","doi":"10.1093/clinchem/hvae106.380","DOIUrl":"https://doi.org/10.1093/clinchem/hvae106.380","url":null,"abstract":"Background ApoAST/ALT, which have lost a coenzyme such as pyridoxal phosphate (PALP), are normally present at low levels in serum. To form the holoenzyme, IFCC recommends adding PALP into AST/ALT reagents. However, commercial reagents often do not use PALP due to its instability. In recent years, pyridoxamine phosphate (PAMP), a more stable coenzyme, has been marketed. However, the differences in holo formation ability and reagent stability between their coenzymes are not well understood. In this study, we focused on the preparation of PAMP and recombinant human ApoAST/ALT (rhApoAST/ALT), followed by fundamental investigations into their use as AST/ALT reagents. Methods PAMP was synthesized from pyridoxamine through an enzymatic reaction using pyridoxal kinase. The reaction product was purified to achieve a purity of >99% in HPLC. rhApoAST/ALT were expressed in Escherichia coli and purified with chromatography. AST/ALT assay was performed on a Hitachi 7180 automatic analyzer. PALP reagents for AST/ALT were designed to correspond with the final concentration of the IFCC method. PAMP reagents were prepared with PAMP instead of PALP. To evaluate the holo formation ability of PALP and PAMP reagents, we measured a dilution series of rhApoAST/ALT. To assess stability, the reagents were stored at 11°C for 3 months and then measured same. The reagents were subjected to content analysis by HPLC. Results Similar results were shown between coenzymes in the evaluation of holo formation ability, and high correlations were observed. As for the stability test, the values of PALP reagents decreased after 3 months, while the values of PAMP reagents did not. Content analysis showed that a spontaneous transamination occurred in PALP reagents. These results were observed at both AST and ALT reagents. Conclusions In AST/ALT reagents, both PAMP and PALP can activate apoenzymes. While PAMP is stable, PALP causes a decrease in reagent performance due to a spontaneous transamination.","PeriodicalId":10690,"journal":{"name":"Clinical chemistry","volume":null,"pages":null},"PeriodicalIF":9.3,"publicationDate":"2024-10-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142362939","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-10-02DOI: 10.1093/clinchem/hvae106.353
B Van Der Pol, R Arcenas, C Boraas, S Chavoustie, L L Crane, N d'Empaire, A C Ermel, G Harnett, F Hinestrosa, S House, R Lillis, J Miller, A Mills, R Poblete, S A Young
Background Screening for Chlamydia trachomatis (CT), Neisseria gonorrhoeae (NG), and Mycoplasma genitalium (MG) can be achieved rapidly (in approximately 20 minutes) with the cobas® CT/NG/MG test (assay not cleared by US FDA. Submission currently under review and subject to change per health authority feedback). This automated, qualitative, real-time PCR-based nucleic acid amplification test (NAAT) is for use on the cobas® liat® system. This study evaluated the test’s clinical performance using urogenital samples from symptomatic and asymptomatic patients. Methods This non-interventional, non-observational study used prospective clinician-collected and self-collected specimens (urine, and vaginal swabs, all in cobas® PCR Media) from symptomatic/asymptomatic patients ≥14 years old from 13 POC sites across the US. The sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of cobas liat CT/NG/MG were calculated with respect to patient infected status (PIS), as determined using results from three FDA-approved NAATs and one laboratory-developed test. Due to insufficient positive NG samples during the trial period, further testing used archived samples. Results The median (range) age of the study population (N=4,800) was 35.0 (15.0-81.0) years; 40.4% (n=1,941) were symptomatic and 51.9% (n=2,489) were assigned female at birth. The cobas CT/NG/MG nucleic acid test demonstrated good clinical performance (Table 1). Across all specimen types, specificity was >97% for each analyte. Sensitivity was ≥95%, except in female urine (CT 87.0%, NG 93.1%, MG 78.9%). Regardless of specimen type, PPV and NPV were ≥95% for CT and NG; PPV for MG was highest in male urine (92.7%) and NPV was >97.5% across analytes. Conclusions In a clinical setting, the cobas CT/NG/MG nucleic acid test showed good clinical performance for the detection of CT, NG, and MG in urogenital samples, in addition to providing a short turn-around time and centralized testing lab quality at the POC for both self- and clinician-collected samples.
{"title":"A-359 Clinical Performance Evaluation of the Polymerase Chain Reaction (PCR)-Based cobas CT/NG/MG Test for Use on the cobas liat System in a Clinical Laboratory Setting and Point-of-Care (POC) Location","authors":"B Van Der Pol, R Arcenas, C Boraas, S Chavoustie, L L Crane, N d'Empaire, A C Ermel, G Harnett, F Hinestrosa, S House, R Lillis, J Miller, A Mills, R Poblete, S A Young","doi":"10.1093/clinchem/hvae106.353","DOIUrl":"https://doi.org/10.1093/clinchem/hvae106.353","url":null,"abstract":"Background Screening for Chlamydia trachomatis (CT), Neisseria gonorrhoeae (NG), and Mycoplasma genitalium (MG) can be achieved rapidly (in approximately 20 minutes) with the cobas® CT/NG/MG test (assay not cleared by US FDA. Submission currently under review and subject to change per health authority feedback). This automated, qualitative, real-time PCR-based nucleic acid amplification test (NAAT) is for use on the cobas® liat® system. This study evaluated the test’s clinical performance using urogenital samples from symptomatic and asymptomatic patients. Methods This non-interventional, non-observational study used prospective clinician-collected and self-collected specimens (urine, and vaginal swabs, all in cobas® PCR Media) from symptomatic/asymptomatic patients ≥14 years old from 13 POC sites across the US. The sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of cobas liat CT/NG/MG were calculated with respect to patient infected status (PIS), as determined using results from three FDA-approved NAATs and one laboratory-developed test. Due to insufficient positive NG samples during the trial period, further testing used archived samples. Results The median (range) age of the study population (N=4,800) was 35.0 (15.0-81.0) years; 40.4% (n=1,941) were symptomatic and 51.9% (n=2,489) were assigned female at birth. The cobas CT/NG/MG nucleic acid test demonstrated good clinical performance (Table 1). Across all specimen types, specificity was >97% for each analyte. Sensitivity was ≥95%, except in female urine (CT 87.0%, NG 93.1%, MG 78.9%). Regardless of specimen type, PPV and NPV were ≥95% for CT and NG; PPV for MG was highest in male urine (92.7%) and NPV was >97.5% across analytes. Conclusions In a clinical setting, the cobas CT/NG/MG nucleic acid test showed good clinical performance for the detection of CT, NG, and MG in urogenital samples, in addition to providing a short turn-around time and centralized testing lab quality at the POC for both self- and clinician-collected samples.","PeriodicalId":10690,"journal":{"name":"Clinical chemistry","volume":null,"pages":null},"PeriodicalIF":9.3,"publicationDate":"2024-10-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142369248","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-10-02DOI: 10.1093/clinchem/hvae106.560
T Jagneaux, A Grantham, K Richard, C Thomas, C D’Antonio, M Laperouse, R Scoggins, H O’Neal
Background In August 2023 Our Lady of the Lake Regional Medical Center implemented a process for sepsis care based on a novel sepsis diagnostic (IntelliSep) in the Emergency Department (ED). As a component of our Sepsis Learning Health Program, we continually evaluate this process. Methods A nurse-driven protocol allows for IntelliSep ordering with triage assessment. Dependent upon bed availability and assessment for clinical stability by the triage staff, patients may be referred to the waiting room after blood draw. An IntelliSep Band 3 result is a critical value, initiating a sepsis pathway. Patients in waiting room at the time of a Band 3 result are immediately placed in an ED bed. We evaluated this process for efficacy and efficiency through review of the medical record. Results Between 01-Sep-2023 and 07-Feb-2024, we performed a total of 2322 IntelliSep tests, with 247 (10.6%) resulting prior to bed assignment, consisting of 124 (50.2%) Band 1, 73 (29.6%) Band 2, and 50 (20.2%) Band 3. The median time to bed assignment (TTB) for these patients was 104 min (Q1-Q3 71-180). TTB for Band 3 (median 73 min, Q1-Q3 58-102) was significantly lower than both Band 1 (146 min, Q1-Q3 86-201 min) and Band 2 (98 min, Q1-Q3 70-174 min), p < 0.0001 and 0.01 respectively. Providers admitted 45 (90%) of Band 3 patients and 62 (50%) of Band 1 patients (p < 0.0001). Discharge diagnosis included infection in 47 (94%) and sepsis in 27 (54%) of Band 3 patients, and infection in 686 (54.8%) and sepsis in 2 (1.6%) of Band 1 patients (p < 0.0001 for infection and sepsis). Conclusions An IntelliSep-based process for sepsis diagnosis, implemented at triage, can expedite identification and treatment of patients presenting to the ED with occult sepsis who appear clinically stable by triage staff.
{"title":"B-200 Expediting Identification of Occult Sepsis with a Novel Diagnostic for Patients Presenting to the ED with Possible Infection","authors":"T Jagneaux, A Grantham, K Richard, C Thomas, C D’Antonio, M Laperouse, R Scoggins, H O’Neal","doi":"10.1093/clinchem/hvae106.560","DOIUrl":"https://doi.org/10.1093/clinchem/hvae106.560","url":null,"abstract":"Background In August 2023 Our Lady of the Lake Regional Medical Center implemented a process for sepsis care based on a novel sepsis diagnostic (IntelliSep) in the Emergency Department (ED). As a component of our Sepsis Learning Health Program, we continually evaluate this process. Methods A nurse-driven protocol allows for IntelliSep ordering with triage assessment. Dependent upon bed availability and assessment for clinical stability by the triage staff, patients may be referred to the waiting room after blood draw. An IntelliSep Band 3 result is a critical value, initiating a sepsis pathway. Patients in waiting room at the time of a Band 3 result are immediately placed in an ED bed. We evaluated this process for efficacy and efficiency through review of the medical record. Results Between 01-Sep-2023 and 07-Feb-2024, we performed a total of 2322 IntelliSep tests, with 247 (10.6%) resulting prior to bed assignment, consisting of 124 (50.2%) Band 1, 73 (29.6%) Band 2, and 50 (20.2%) Band 3. The median time to bed assignment (TTB) for these patients was 104 min (Q1-Q3 71-180). TTB for Band 3 (median 73 min, Q1-Q3 58-102) was significantly lower than both Band 1 (146 min, Q1-Q3 86-201 min) and Band 2 (98 min, Q1-Q3 70-174 min), p &lt; 0.0001 and 0.01 respectively. Providers admitted 45 (90%) of Band 3 patients and 62 (50%) of Band 1 patients (p &lt; 0.0001). Discharge diagnosis included infection in 47 (94%) and sepsis in 27 (54%) of Band 3 patients, and infection in 686 (54.8%) and sepsis in 2 (1.6%) of Band 1 patients (p &lt; 0.0001 for infection and sepsis). Conclusions An IntelliSep-based process for sepsis diagnosis, implemented at triage, can expedite identification and treatment of patients presenting to the ED with occult sepsis who appear clinically stable by triage staff.","PeriodicalId":10690,"journal":{"name":"Clinical chemistry","volume":null,"pages":null},"PeriodicalIF":9.3,"publicationDate":"2024-10-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142369004","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-10-02DOI: 10.1093/clinchem/hvae106.528
Y Xiao, M Wakefield, M Gabra, E Leung
Background Urine organic acid (UOA) analysis is essential for the diagnosis of inborn errors of metabolism (IEMs). Traditionally, UOA analysis is performed with gas chromatography-mass spectrometry (GC-MS) and requires time-consuming sample preparation steps including liquid-liquid extraction and derivatization. The rapid development of Ultra-Performance Liquid Chromatography Tandem Mass Spectrometry (UPLC-MS/MS) in the past few years provides the opportunity to perform UOA analysis with a dilute-and-shoot methodology. We describe the development and validation of a quantitative Ultra-Performance Liquid Chromatography Quadrupole Time-of-Flight (UPLC-QToF) method for UOA analysis. Methods Urine specimens were diluted to normalize creatinine concentrations to 1 mmol/L. 20 µL of urine specimen (diluted), calibrator, or quality control (QC) material was mixed with 400 µL of mobile phase A (0.05% formic acid in water) and a mixture of isotope-labeled internal standards. After centrifugation, 10 µL of the supernatant was analyzed using a Xevo G3 QTOF mass spectrometer (Waters) with a ACQUITYTM Premier HSS T3 1.8 µm VanGuardTM FIT 2.1 x 150 mm column (Waters). Data collection was performed with negative electrospray ionization (ESI) mode using the MSE method to produce fragment ions when applicable. Repeatability, reproducibility, and carryover were assessed using the QC materials. The analytical measuring range (AMR) was assessed using synthetic urine spiked with increasing concentrations of each organic acid. Accuracy was assessed by method comparison with the UOA test performed at Mayo Clinic Laboratory and by spike-recovery study using a pooled urine specimen. Matrix effect was also evaluated with matrix dilution study. Results An optimized LC method was used to enable high-resolution separation of selected UOAs (N = 29) and isomers. Total analytical time was 20 min per injection. Both linear and quadratic regressions were used to build the calibration curves. AMR and correlation coefficients of a few representative UOAs were: orotic acid (3.4 to 214.2 mmol/mol creatinine, R^2 = 0.99, linear regression); 2-methylcitric acid (4 to 189 mmol/mol creatinine, R^2 = 0.99, linear regression); 3-methylcrotonylglycine (0.3 to 18.0 mmol/mol creatinine, R^2 = 0.99, linear regression). Repeatability and reproducibility were mostly <=10% CV and no carryover was observed. Spike-recovery study demonstrated recoveries between 80% and 120%, and method comparison study demonstrated no discrepancies with results from Mayo Clinic Laboratory. Conclusions We have developed and validated a novel UPLC-QTOF method for UOA analysis to support the diagnosis of IEMs with acceptable analytical and clinical performances. Compared with the traditional GC-MS method, the UPLC-QTOF method requires a very small specimen volume and does not require laborious and time-consuming sample preparation steps. Continued optimization of the method will be pursued to measure more UOAs to
{"title":"B-168 Development and Validation of a Quantitative Ultra-Performance Liquid Chromatography Quadrupole Time-of-Flight (UPLC-QToF) Method for Urine Organic Acid Analysis","authors":"Y Xiao, M Wakefield, M Gabra, E Leung","doi":"10.1093/clinchem/hvae106.528","DOIUrl":"https://doi.org/10.1093/clinchem/hvae106.528","url":null,"abstract":"Background Urine organic acid (UOA) analysis is essential for the diagnosis of inborn errors of metabolism (IEMs). Traditionally, UOA analysis is performed with gas chromatography-mass spectrometry (GC-MS) and requires time-consuming sample preparation steps including liquid-liquid extraction and derivatization. The rapid development of Ultra-Performance Liquid Chromatography Tandem Mass Spectrometry (UPLC-MS/MS) in the past few years provides the opportunity to perform UOA analysis with a dilute-and-shoot methodology. We describe the development and validation of a quantitative Ultra-Performance Liquid Chromatography Quadrupole Time-of-Flight (UPLC-QToF) method for UOA analysis. Methods Urine specimens were diluted to normalize creatinine concentrations to 1 mmol/L. 20 µL of urine specimen (diluted), calibrator, or quality control (QC) material was mixed with 400 µL of mobile phase A (0.05% formic acid in water) and a mixture of isotope-labeled internal standards. After centrifugation, 10 µL of the supernatant was analyzed using a Xevo G3 QTOF mass spectrometer (Waters) with a ACQUITYTM Premier HSS T3 1.8 µm VanGuardTM FIT 2.1 x 150 mm column (Waters). Data collection was performed with negative electrospray ionization (ESI) mode using the MSE method to produce fragment ions when applicable. Repeatability, reproducibility, and carryover were assessed using the QC materials. The analytical measuring range (AMR) was assessed using synthetic urine spiked with increasing concentrations of each organic acid. Accuracy was assessed by method comparison with the UOA test performed at Mayo Clinic Laboratory and by spike-recovery study using a pooled urine specimen. Matrix effect was also evaluated with matrix dilution study. Results An optimized LC method was used to enable high-resolution separation of selected UOAs (N = 29) and isomers. Total analytical time was 20 min per injection. Both linear and quadratic regressions were used to build the calibration curves. AMR and correlation coefficients of a few representative UOAs were: orotic acid (3.4 to 214.2 mmol/mol creatinine, R^2 = 0.99, linear regression); 2-methylcitric acid (4 to 189 mmol/mol creatinine, R^2 = 0.99, linear regression); 3-methylcrotonylglycine (0.3 to 18.0 mmol/mol creatinine, R^2 = 0.99, linear regression). Repeatability and reproducibility were mostly &lt;=10% CV and no carryover was observed. Spike-recovery study demonstrated recoveries between 80% and 120%, and method comparison study demonstrated no discrepancies with results from Mayo Clinic Laboratory. Conclusions We have developed and validated a novel UPLC-QTOF method for UOA analysis to support the diagnosis of IEMs with acceptable analytical and clinical performances. Compared with the traditional GC-MS method, the UPLC-QTOF method requires a very small specimen volume and does not require laborious and time-consuming sample preparation steps. Continued optimization of the method will be pursued to measure more UOAs to","PeriodicalId":10690,"journal":{"name":"Clinical chemistry","volume":null,"pages":null},"PeriodicalIF":9.3,"publicationDate":"2024-10-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142369121","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-10-02DOI: 10.1093/clinchem/hvae106.087
L M Leonard, M A Nicklas, D R Block, N A Baumann
Background Thoroughly investigating single patient result errors may identify systemic, large-scale testing errors. The laboratory observed an event where a revision to a Total Bile Acid (TBA, Diazyme Laboratories, Inc.) patient result following a 22S quality control (QC) failure displayed a larger than expected difference (52 to 5mcmol/L). TBA is primarily used for diagnosis and monitoring of intrahepatic cholestasis of pregnancy with results >10mcmol/L considered elevated. Retrospective review of result revisions following QC failures (3/15/2017-5/30/2023) revealed that 50%(20/40) yielded result differences of >10mcmol/L(>20SD based on assay imprecision). The aim of this study was to apply a systematic approach to i) estimate the rate of TBA errors, ii) ensure accurate result reporting during investigation period, iii) perform root cause analysis (RCA), and iv) determine corrective and preventative action(s). Methods Residual samples from TBA testing (6/24-7/5/2023) were retested(n=158), differences >+/-3.0mcmol/L or 15% were confirmed on an alternate analyzer, and reports were revised. Automated repeat testing of TBA samples >10mcmol/L was operationalized 7/6/2023. Initial and repeat TBA results were compared(n=448) and results differing by >+/-20% were remeasured on an alternate analyzer. RCA was conducted using a fishbone diagram. NaOH reagent probe washes were implemented for TBA and the error rate was re-assessed. Assays run prior to TBA samples with errors (n=15) were identified. To assess reagent carryover a residual serum pool (TBA ∼5mcmol/L) was aliquoted into 5 tubes in a sample rack. The first sample in the rack was programmed to run an assay suspected of causing carryover followed by 4 TBA measurements. Mean±standard deviation(SD) TBA concentrations were calculated. TBA was also measured in the liquid reagent for amylase, lipase, acetylcholinesterase (ACE) and fructosamine. Results Initial TBA retesting yielded no errors when initial TBA was ≤10mcmol/L(n=51). For samples with TBA >10mcmol/L(n=107), 9(8.4%) had differences exceeding criteria with 8/9 being revised to ≤10mcmol/L. Analysis of automated patient repeat data showed a 3.8%(17/448) error rate when initial TBA >10mcmol/L. After NaOH reagent probe washes were implemented, the error rate decreased to 0.7%(3/448). Assays run directly before an erroneously high TBA result included: lactate, fructosamine, soluble transferrin receptor(STFR), lipase, and ACE. A serum TBA pool(mean±SD=5.1±0.4 mcmol/L,n=15) measured 37.6±1.2mcmol/L(n=3) and 6.7±0.7mcmol/L(n=3) after lipase and fructosamine, respectively. No other assays demonstrated carryover. TBA in lipase reagent compartment B and C was 653 and 649mcmol/L, respectively, and 653mcmol/L in fructosamine reagent compartment B. Conclusions A single patient TBA result revision was investigated and led to identification of reagent carryover causing erroneously high TBA results on the
{"title":"A-088 Clinically Significant Errors Using the Diazyme Total Bile Acid Assay on the Roche c502 Analyzer: Investigating a Result Error Identifies a Novel Large-Scale Testing Error Caused by Reagent Carryover","authors":"L M Leonard, M A Nicklas, D R Block, N A Baumann","doi":"10.1093/clinchem/hvae106.087","DOIUrl":"https://doi.org/10.1093/clinchem/hvae106.087","url":null,"abstract":"Background Thoroughly investigating single patient result errors may identify systemic, large-scale testing errors. The laboratory observed an event where a revision to a Total Bile Acid (TBA, Diazyme Laboratories, Inc.) patient result following a 22S quality control (QC) failure displayed a larger than expected difference (52 to 5mcmol/L). TBA is primarily used for diagnosis and monitoring of intrahepatic cholestasis of pregnancy with results &gt;10mcmol/L considered elevated. Retrospective review of result revisions following QC failures (3/15/2017-5/30/2023) revealed that 50%(20/40) yielded result differences of &gt;10mcmol/L(&gt;20SD based on assay imprecision). The aim of this study was to apply a systematic approach to i) estimate the rate of TBA errors, ii) ensure accurate result reporting during investigation period, iii) perform root cause analysis (RCA), and iv) determine corrective and preventative action(s). Methods Residual samples from TBA testing (6/24-7/5/2023) were retested(n=158), differences &gt;+/-3.0mcmol/L or 15% were confirmed on an alternate analyzer, and reports were revised. Automated repeat testing of TBA samples &gt;10mcmol/L was operationalized 7/6/2023. Initial and repeat TBA results were compared(n=448) and results differing by &gt;+/-20% were remeasured on an alternate analyzer. RCA was conducted using a fishbone diagram. NaOH reagent probe washes were implemented for TBA and the error rate was re-assessed. Assays run prior to TBA samples with errors (n=15) were identified. To assess reagent carryover a residual serum pool (TBA ∼5mcmol/L) was aliquoted into 5 tubes in a sample rack. The first sample in the rack was programmed to run an assay suspected of causing carryover followed by 4 TBA measurements. Mean±standard deviation(SD) TBA concentrations were calculated. TBA was also measured in the liquid reagent for amylase, lipase, acetylcholinesterase (ACE) and fructosamine. Results Initial TBA retesting yielded no errors when initial TBA was ≤10mcmol/L(n=51). For samples with TBA &gt;10mcmol/L(n=107), 9(8.4%) had differences exceeding criteria with 8/9 being revised to ≤10mcmol/L. Analysis of automated patient repeat data showed a 3.8%(17/448) error rate when initial TBA &gt;10mcmol/L. After NaOH reagent probe washes were implemented, the error rate decreased to 0.7%(3/448). Assays run directly before an erroneously high TBA result included: lactate, fructosamine, soluble transferrin receptor(STFR), lipase, and ACE. A serum TBA pool(mean±SD=5.1±0.4 mcmol/L,n=15) measured 37.6±1.2mcmol/L(n=3) and 6.7±0.7mcmol/L(n=3) after lipase and fructosamine, respectively. No other assays demonstrated carryover. TBA in lipase reagent compartment B and C was 653 and 649mcmol/L, respectively, and 653mcmol/L in fructosamine reagent compartment B. Conclusions A single patient TBA result revision was investigated and led to identification of reagent carryover causing erroneously high TBA results on the","PeriodicalId":10690,"journal":{"name":"Clinical chemistry","volume":null,"pages":null},"PeriodicalIF":9.3,"publicationDate":"2024-10-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142369123","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-10-02DOI: 10.1093/clinchem/hvae106.480
J Choi
Background Copy Number Variations (CNVs) are critical genetic markers in diversity and disease, yet their accurate extraction from medical literature remains challenging due to the complexity of genetic data. While specialized NLP models like CNV-ETLAI have been developed for this task, the advent of Large Language Models (LLMs) such as GPT-4 presents a potential alternative with broader applicability. This study evaluates the efficacy of GPT-4 against CNV-ETLAI in extracting CNVs from medical journal articles, aiming to enhance genetic research and clinical decision-making. Methods We configured GPT-4 to process and interpret medical journal PDFs, designing custom prompts for CNV information extraction. The performance of GPT-4 was benchmarked against CNV-ETLAI using a dataset of 146 true positive CNVs extracted from 23 journal articles. Performance metrics focused on accuracy in extracting CNVs from both text and tables, recognizing the importance of structured data interpretation in genomic analysis. Results CNV-ETLAI demonstrated superior accuracy, achieving a 98% success rate in CNV extraction, compared to GPT-4’s 49%. Specifically, CNV-ETLAI outperformed GPT-4 in table extraction accuracy (99% vs. 41.2%) and context extraction accuracy (96% vs. 63.2%). Despite GPT-4's lower performance, its capacity for improvement and adaptability was noted, indicating potential future applicability in medical data extraction. Conclusions The study highlights CNV-ETLAI's current superiority in extracting CNVs from medical texts, particularly in interpreting structured data like tables. However, the adaptability and potential for growth in LLMs like GPT-4 suggest they could soon become valuable tools for medical data extraction, offering a more versatile and powerful solution across a broader range of applications. The promise of LLMs, despite their current limitations, underscores the need for continued research and development in AI technologies for genomic data interpretation.
{"title":"B-119 Comparative performance of GPT-4 and CNV-ETLAI in extracting copy number variations from medical journals: Bridging the gap between large language models and specialized NLP tools in genomic data interpretation","authors":"J Choi","doi":"10.1093/clinchem/hvae106.480","DOIUrl":"https://doi.org/10.1093/clinchem/hvae106.480","url":null,"abstract":"Background Copy Number Variations (CNVs) are critical genetic markers in diversity and disease, yet their accurate extraction from medical literature remains challenging due to the complexity of genetic data. While specialized NLP models like CNV-ETLAI have been developed for this task, the advent of Large Language Models (LLMs) such as GPT-4 presents a potential alternative with broader applicability. This study evaluates the efficacy of GPT-4 against CNV-ETLAI in extracting CNVs from medical journal articles, aiming to enhance genetic research and clinical decision-making. Methods We configured GPT-4 to process and interpret medical journal PDFs, designing custom prompts for CNV information extraction. The performance of GPT-4 was benchmarked against CNV-ETLAI using a dataset of 146 true positive CNVs extracted from 23 journal articles. Performance metrics focused on accuracy in extracting CNVs from both text and tables, recognizing the importance of structured data interpretation in genomic analysis. Results CNV-ETLAI demonstrated superior accuracy, achieving a 98% success rate in CNV extraction, compared to GPT-4’s 49%. Specifically, CNV-ETLAI outperformed GPT-4 in table extraction accuracy (99% vs. 41.2%) and context extraction accuracy (96% vs. 63.2%). Despite GPT-4's lower performance, its capacity for improvement and adaptability was noted, indicating potential future applicability in medical data extraction. Conclusions The study highlights CNV-ETLAI's current superiority in extracting CNVs from medical texts, particularly in interpreting structured data like tables. However, the adaptability and potential for growth in LLMs like GPT-4 suggest they could soon become valuable tools for medical data extraction, offering a more versatile and powerful solution across a broader range of applications. The promise of LLMs, despite their current limitations, underscores the need for continued research and development in AI technologies for genomic data interpretation.","PeriodicalId":10690,"journal":{"name":"Clinical chemistry","volume":null,"pages":null},"PeriodicalIF":9.3,"publicationDate":"2024-10-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142369047","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-10-02DOI: 10.1093/clinchem/hvae106.082
S Brambilla, C Assalini, E Frana, R Lucini, D Novati
Background The analytical performances of the Angiotensin Converting Enzyme (ACE) assay for quantitative determination of ACE activity in human serum and plasma have been evaluated on the Abbott Alinity c system and ARCHITECT c16000 System. ACE (dipeptidyl carboxypeptidase) is a glycoprotein peptidyldipeptide hydrolase that cleaves histidylleucine dipeptide from angiotensin I, a relatively inactive decapeptide. The latter is converted to the potent vasoconstrictor, angiotensin II. ACE also inactivates bradykinin. Elevated levels of ACE activity occur in serum of patients with active sarcoidosis, and occasionally in premature infants with respiratory distress syndrome, in adults with tuberculosis, Gaucher’s disease, leprosy, and in many other pathologic conditions involving lung and liver diseases. Methods ACE hydrolyses urylacryloylphenylalanine-glycylglycine (FAPGG) to furylacryloylphenylalanine (FAP) and glycylglycine. Hydrolysis of FAPGG results in a decrease in absorbance at 340 nm. The ACE activity in the sample is determined by comparing the sample reaction rate to that obtained with the Sentinel ACE calibrator. Performances evaluation have been done following the latest CLSI guidelines protocols available. Results Summary of the analytical performances for the tested assay is reported in table below. Conclusions Analytical performances of Sentinel Diagnostics’ ACE assay demonstrate that quantitative determination of ACE activity in human serum and plasma are suitable for the routine measurement of the analyte on Abbott Alinity c system and ARCHITECT c16000 System.
{"title":"A-083 Performance evaluation of Sentinel Diagnostics’ ACE assay on the Abbott Alinity c system and ARCHITECT c16000 System","authors":"S Brambilla, C Assalini, E Frana, R Lucini, D Novati","doi":"10.1093/clinchem/hvae106.082","DOIUrl":"https://doi.org/10.1093/clinchem/hvae106.082","url":null,"abstract":"Background The analytical performances of the Angiotensin Converting Enzyme (ACE) assay for quantitative determination of ACE activity in human serum and plasma have been evaluated on the Abbott Alinity c system and ARCHITECT c16000 System. ACE (dipeptidyl carboxypeptidase) is a glycoprotein peptidyldipeptide hydrolase that cleaves histidylleucine dipeptide from angiotensin I, a relatively inactive decapeptide. The latter is converted to the potent vasoconstrictor, angiotensin II. ACE also inactivates bradykinin. Elevated levels of ACE activity occur in serum of patients with active sarcoidosis, and occasionally in premature infants with respiratory distress syndrome, in adults with tuberculosis, Gaucher’s disease, leprosy, and in many other pathologic conditions involving lung and liver diseases. Methods ACE hydrolyses urylacryloylphenylalanine-glycylglycine (FAPGG) to furylacryloylphenylalanine (FAP) and glycylglycine. Hydrolysis of FAPGG results in a decrease in absorbance at 340 nm. The ACE activity in the sample is determined by comparing the sample reaction rate to that obtained with the Sentinel ACE calibrator. Performances evaluation have been done following the latest CLSI guidelines protocols available. Results Summary of the analytical performances for the tested assay is reported in table below. Conclusions Analytical performances of Sentinel Diagnostics’ ACE assay demonstrate that quantitative determination of ACE activity in human serum and plasma are suitable for the routine measurement of the analyte on Abbott Alinity c system and ARCHITECT c16000 System.","PeriodicalId":10690,"journal":{"name":"Clinical chemistry","volume":null,"pages":null},"PeriodicalIF":9.3,"publicationDate":"2024-10-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142368788","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-10-02DOI: 10.1093/clinchem/hvae106.203
E Jurinic
Background Recent pandemics, epidemics and outbreaks continually underscore the critical importance of resilient supply chains in healthcare, particularly in clinical laboratories where timely access to supplies is essential for patient care. Supply chain disruptions, whether due to global crises or localized challenges, can significantly impact laboratory operations, leading to delayed diagnoses, compromised patient outcomes, and increased costs. It is estimated that 70%-80% of a patients’ EMR is clinical laboratory results. Despite this, supply chain resilience in clinical laboratories remains an under-addressed issue and remains a buzzword. Methods To evaluate the cost and importance of supply chain resilience in the clinical laboratory, we conducted a comprehensive review and interviews with 9 Healthcare Supply Chain experts analyzing the impact of supply chain disruptions on laboratory operations and patient care. Data was also collected from a diverse range of clinical laboratories, including hospitals, independent labs, and research institutions, spanning different geographic regions as well as a survey of 50 Laboratory experts with a 50% response rate. Key metrics assessed included: -Frequency and duration of supply chain disruptions. -Financial costs incurred due to disruptions, including expenses related to expedited shipping, alternative sourcing, and inventory management. -Impact on laboratory workflow, turnaround times, and patient care outcomes. -Strategies employed to mitigate supply chain risks and enhance resilience. Results Our review revealed findings regarding the cost and importance of supply chain resilience in the clinical laboratory: -Supply chain disruptions were frequent, with 85% of laboratories experiencing at least one significant disruption in the past year and 100% in experienced disruptions 2022. -The average duration of disruptions was 2-3 weeks, leading to substantial delays in test processing and patient care. -Financial costs associated with disruptions averaged $50,000 per laboratory per annum, including expenses for rush orders, and premium-priced alternatives. -Common strategies employed were increased inventory or safety stocks, multi-vendor sourcing, and supply chain mapping. Conclusions The findings of this study underscore the critical importance of supply chain resilience in the clinical laboratory. Supply chain disruptions are not just inconveniences; they pose significant risks to patient safety, operational efficiency, and financial sustainability. Investing in supply chain resilience is imperative for laboratories to fulfill their mission of delivering timely and accurate patient results. To continue to enhance supply chain resilience, laboratories must adopt a proactive approach, leveraging data-driven insights and collaborative partnerships. This includes diversifying supplier networks and fostering closer collaboration between laboratory and procurement teams.
{"title":"A-205 Evaluating the Cost and Importance of Supply Chain Resilience in the Clinical Laboratory","authors":"E Jurinic","doi":"10.1093/clinchem/hvae106.203","DOIUrl":"https://doi.org/10.1093/clinchem/hvae106.203","url":null,"abstract":"Background Recent pandemics, epidemics and outbreaks continually underscore the critical importance of resilient supply chains in healthcare, particularly in clinical laboratories where timely access to supplies is essential for patient care. Supply chain disruptions, whether due to global crises or localized challenges, can significantly impact laboratory operations, leading to delayed diagnoses, compromised patient outcomes, and increased costs. It is estimated that 70%-80% of a patients’ EMR is clinical laboratory results. Despite this, supply chain resilience in clinical laboratories remains an under-addressed issue and remains a buzzword. Methods To evaluate the cost and importance of supply chain resilience in the clinical laboratory, we conducted a comprehensive review and interviews with 9 Healthcare Supply Chain experts analyzing the impact of supply chain disruptions on laboratory operations and patient care. Data was also collected from a diverse range of clinical laboratories, including hospitals, independent labs, and research institutions, spanning different geographic regions as well as a survey of 50 Laboratory experts with a 50% response rate. Key metrics assessed included: -Frequency and duration of supply chain disruptions. -Financial costs incurred due to disruptions, including expenses related to expedited shipping, alternative sourcing, and inventory management. -Impact on laboratory workflow, turnaround times, and patient care outcomes. -Strategies employed to mitigate supply chain risks and enhance resilience. Results Our review revealed findings regarding the cost and importance of supply chain resilience in the clinical laboratory: -Supply chain disruptions were frequent, with 85% of laboratories experiencing at least one significant disruption in the past year and 100% in experienced disruptions 2022. -The average duration of disruptions was 2-3 weeks, leading to substantial delays in test processing and patient care. -Financial costs associated with disruptions averaged $50,000 per laboratory per annum, including expenses for rush orders, and premium-priced alternatives. -Common strategies employed were increased inventory or safety stocks, multi-vendor sourcing, and supply chain mapping. Conclusions The findings of this study underscore the critical importance of supply chain resilience in the clinical laboratory. Supply chain disruptions are not just inconveniences; they pose significant risks to patient safety, operational efficiency, and financial sustainability. Investing in supply chain resilience is imperative for laboratories to fulfill their mission of delivering timely and accurate patient results. To continue to enhance supply chain resilience, laboratories must adopt a proactive approach, leveraging data-driven insights and collaborative partnerships. This includes diversifying supplier networks and fostering closer collaboration between laboratory and procurement teams.","PeriodicalId":10690,"journal":{"name":"Clinical chemistry","volume":null,"pages":null},"PeriodicalIF":9.3,"publicationDate":"2024-10-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142369119","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-10-02DOI: 10.1093/clinchem/hvae106.619
M Bohn, S Delaney, B Jung, W Lamb, F Leung
Background Point-of-care (POC) urine drug testing is a useful adjunct to self-reporting in rapid access addiction medicine settings to immediately guide patient management. However, available POC immunoassays have limitations including cross-reactivity with unrelated compounds or low sensitivity that may cause false results. Here, we assessed the performance of a multi-drug test panel by comparing against gold-standard liquid chromatography tandem mass spectrometry (LC-MS/MS) testing. Methods 102 residual urine specimens were assayed using a competitive lateral flow immunoassay (LFA) for 10 drugs: 2-ethylene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP, methadone metabolite), buprenorphine, morphine, hydromorphone, oxycodone, fentanyl, cocaine, methamphetamine, amphetamine, and benzodiazepines (BTNX Rapid ResponseTM Multi-Drug Panel). Results were compared to those obtained by LC-MS/MS (n=67, 66%) or kinetic interaction of microparticles in solution automated immunoassay (KIMS) (Roche Diagnostics, n=35, 33%). Broad spectrum LC-MS/MS results were reviewed in entirety for discordant cases, particularly in false positives to identify the presence of any known cross-reacting compounds. Results Of 10 drugs evaluated, four demonstrated ≥95% agreement with LC-MS/MS or KIMS immunoassay (EDDP, buprenorphine, oxycodone, methamphetamine). Fentanyl demonstrated the highest false negative rate of 44% (LFA cut-off: 10 ng/mL) followed by amphetamines (22%, cut-off: 1000 ng/mL). Morphine and hydromorphone demonstrated false positive rates of 14% and 18%, respectively, with most cases likely due to cross-reacting opiate or opioid metabolites. Benzodiazepines (target: Oxazepam) demonstrated false positive and negative rates of 13% and 24%, respectively. Conclusions To our knowledge, this is the first study to evaluate the performance of the BTNX multi-drug test panel against definitive testing in urine samples. While good concordance was observed for most drugs, high rates of discordant results for fentanyl and others emphasize the need for confirmatory testing by methods with higher sensitivity and specificity. Careful consideration prior to implementation would be essential, including physician education, interpretative comments, and training resources.
{"title":"B-262 Validation of a point-of-care lateral flow immunoassay for urine drug testing prior to application in an outpatient rapid access addiction medicine clinic","authors":"M Bohn, S Delaney, B Jung, W Lamb, F Leung","doi":"10.1093/clinchem/hvae106.619","DOIUrl":"https://doi.org/10.1093/clinchem/hvae106.619","url":null,"abstract":"Background Point-of-care (POC) urine drug testing is a useful adjunct to self-reporting in rapid access addiction medicine settings to immediately guide patient management. However, available POC immunoassays have limitations including cross-reactivity with unrelated compounds or low sensitivity that may cause false results. Here, we assessed the performance of a multi-drug test panel by comparing against gold-standard liquid chromatography tandem mass spectrometry (LC-MS/MS) testing. Methods 102 residual urine specimens were assayed using a competitive lateral flow immunoassay (LFA) for 10 drugs: 2-ethylene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP, methadone metabolite), buprenorphine, morphine, hydromorphone, oxycodone, fentanyl, cocaine, methamphetamine, amphetamine, and benzodiazepines (BTNX Rapid ResponseTM Multi-Drug Panel). Results were compared to those obtained by LC-MS/MS (n=67, 66%) or kinetic interaction of microparticles in solution automated immunoassay (KIMS) (Roche Diagnostics, n=35, 33%). Broad spectrum LC-MS/MS results were reviewed in entirety for discordant cases, particularly in false positives to identify the presence of any known cross-reacting compounds. Results Of 10 drugs evaluated, four demonstrated ≥95% agreement with LC-MS/MS or KIMS immunoassay (EDDP, buprenorphine, oxycodone, methamphetamine). Fentanyl demonstrated the highest false negative rate of 44% (LFA cut-off: 10 ng/mL) followed by amphetamines (22%, cut-off: 1000 ng/mL). Morphine and hydromorphone demonstrated false positive rates of 14% and 18%, respectively, with most cases likely due to cross-reacting opiate or opioid metabolites. Benzodiazepines (target: Oxazepam) demonstrated false positive and negative rates of 13% and 24%, respectively. Conclusions To our knowledge, this is the first study to evaluate the performance of the BTNX multi-drug test panel against definitive testing in urine samples. While good concordance was observed for most drugs, high rates of discordant results for fentanyl and others emphasize the need for confirmatory testing by methods with higher sensitivity and specificity. Careful consideration prior to implementation would be essential, including physician education, interpretative comments, and training resources.","PeriodicalId":10690,"journal":{"name":"Clinical chemistry","volume":null,"pages":null},"PeriodicalIF":9.3,"publicationDate":"2024-10-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142369175","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-10-02DOI: 10.1093/clinchem/hvae106.539
H Kalil, S Maher, M Bayachou
Background Cisplatin [cis-dichlorodiamine platinum (II)], is a well-recognized chemotherapeutical drug. Cisplatin has been employed in treating a wide range of human cancers, such as those of the breast, bladder, lung, ovarian, and testicular cancers. Its therapeutic action is attributed to its capacity to form crosslinks with the DNA's purine bases, disrupting DNA repair processes, causing DNA damage, and ultimately leading to apoptosis in cancerous cells. Nonetheless, the application of cisplatin is constrained by the development of multidrug resistance and the occurrence of severe adverse effects, including nephrotoxicity, bone marrow suppression, hearing loss, allergic responses, nerve damage, and low magnesium levels in the blood. Numerous strategies have been explored to enhance the anticancer effectiveness of cisplatin while minimizing its associated toxicities. These strategies include combination therapies that incorporate nanoparticles, liposomes, and polymer micelles. In this project, cisplatin was embedded into alginate hydrogels loaded with silver nanoparticles and in vitro cisplatin release study using HPLC and UV-Vis spectrophotometry were studied. Methods We developed a novel nanocomplex by integrating silver nanoparticles (AgNPs) with alginate hydrogel coating to create a versatile platform for drug delivery. To incorporate the chemotherapeutic agent, cisplatin (150 ppm) was introduced into the AgNPs-alginate mixture utilizing rapid stirring to ensure uniform distribution and encapsulation of cisplatin within the nanocomplex. A range of analytical methods, such as UV-Vis, FTIR, SEM/EDX, and Zeta potential analysis, were employed to characterize the nanocomplex. To evaluate cisplatin release kinetics from the nanocomplex, we employed an in vitro dialysis method for monitoring its sustained release. High-performance liquid chromatography (HPLC) was used for precise cisplatin release quantification, then validated by colorimetric spectrophotometry. This dual-method approach ensured accurate insights into the nanocomplex’s release dynamics, substantiating its potential in enhancing targeted cancer therapy through advanced drug delivery systems. Results The analysis through UV-Vis revealed an absorption spectrum around 410 nm, indicative of the characteristic plasmon resonance associated with silver nanoparticles. TEM provided high-resolution imagery, revealing that the size of the silver nanoparticles varied between 4 and 30 nm, averaging at 13 nm with a standard deviation of 5 nm specifically for the alginate-coated AgNPs. SEM images confirmed the anticipated spherical distribution of silver nanoparticles within the alginate hydrogel framework. EDX spectroscopy analysis further verified the incorporation of silver nanoparticles and platinum within the complex. The cisplatin release studies from this newly developed nanocomplex illustrated a prolonged, slow, and consistent release pattern, extending over days, in stark contrast to th
背景 顺铂[顺式二氯二胺铂（II）]是一种公认的化疗药物。顺铂已被用于治疗多种人类癌症，如乳腺癌、膀胱癌、肺癌、卵巢癌和睾丸癌。其治疗作用归因于它能与 DNA 的嘌呤碱基形成交联，破坏 DNA 修复过程，造成 DNA 损伤，最终导致癌细胞凋亡。然而，顺铂的应用受到多药耐药性和严重不良反应的限制，这些不良反应包括肾毒性、骨髓抑制、听力损失、过敏反应、神经损伤和血液中镁含量过低。为了提高顺铂的抗癌效果，同时最大限度地减少其相关毒性，人们探索了许多策略。这些策略包括结合纳米粒子、脂质体和聚合物胶束的组合疗法。在本项目中，我们将顺铂嵌入装有银纳米粒子的藻酸盐水凝胶中，并使用高效液相色谱法和紫外可见分光光度法对顺铂的体外释放进行了研究。方法 我们将银纳米粒子（AgNPs）与海藻酸盐水凝胶涂层相结合，开发出了一种新型纳米复合物，从而创建了一个多功能的给药平台。为了加入化疗药物，我们利用快速搅拌将顺铂（150 ppm）引入 AgNPs-海藻酸盐混合物，以确保顺铂在纳米复合物中的均匀分布和包裹。采用了一系列分析方法，如紫外可见光、傅立叶变换红外光谱、扫描电镜/电子显微镜和 Zeta 电位分析，对纳米复合物进行了表征。为了评估顺铂从纳米复合物中的释放动力学，我们采用了体外透析法来监测其持续释放。采用高效液相色谱法（HPLC）对顺铂释放进行精确定量，然后用比色分光光度法进行验证。这种双重方法确保了对纳米复合物释放动态的准确了解，证实了其通过先进的给药系统加强癌症靶向治疗的潜力。结果 紫外可见光谱分析显示了 410 纳米附近的吸收光谱，表明了与银纳米粒子相关的等离子共振特征。TEM 提供了高分辨率图像，显示银纳米粒子的大小在 4 纳米到 30 纳米之间，平均为 13 纳米，标准偏差为 5 纳米，特别是藻酸盐包裹的 AgNPs。SEM 图像证实了银纳米粒子在海藻酸盐水凝胶框架内的预期球形分布。EDX 光谱分析进一步证实了银纳米粒子和铂在复合物中的结合。对这种新开发的纳米复合物进行的顺铂释放研究表明，其释放模式持久、缓慢且稳定，可持续数天，这与顺铂在未结合状态下的快速完全释放形成了鲜明对比，后者在一小时内就达到了释放峰值。结论 在本研究中，我们成功开发并验证了一种创新的多功能纳米给药平台，该平台由协同共载银纳米粒子（AgNPs）和顺铂的藻酸盐水凝胶组成。这种新型药物载体系统明显增强了顺铂的控制给药能力，三天内缓慢而持续的释放曲线就是证明。与游离态相比，海藻酸盐复合物能缓慢而持续地释放顺铂，这凸显了纳米平台在调节药物释放动力学方面的功效。
{"title":"B-179 Comparative Evaluation of Cisplatin Release from Alginate Hydrogels with Embedded Silver Nanoparticles: An HPLC and Colorimetric Spectrophotometry Study","authors":"H Kalil, S Maher, M Bayachou","doi":"10.1093/clinchem/hvae106.539","DOIUrl":"https://doi.org/10.1093/clinchem/hvae106.539","url":null,"abstract":"Background Cisplatin [cis-dichlorodiamine platinum (II)], is a well-recognized chemotherapeutical drug. Cisplatin has been employed in treating a wide range of human cancers, such as those of the breast, bladder, lung, ovarian, and testicular cancers. Its therapeutic action is attributed to its capacity to form crosslinks with the DNA's purine bases, disrupting DNA repair processes, causing DNA damage, and ultimately leading to apoptosis in cancerous cells. Nonetheless, the application of cisplatin is constrained by the development of multidrug resistance and the occurrence of severe adverse effects, including nephrotoxicity, bone marrow suppression, hearing loss, allergic responses, nerve damage, and low magnesium levels in the blood. Numerous strategies have been explored to enhance the anticancer effectiveness of cisplatin while minimizing its associated toxicities. These strategies include combination therapies that incorporate nanoparticles, liposomes, and polymer micelles. In this project, cisplatin was embedded into alginate hydrogels loaded with silver nanoparticles and in vitro cisplatin release study using HPLC and UV-Vis spectrophotometry were studied. Methods We developed a novel nanocomplex by integrating silver nanoparticles (AgNPs) with alginate hydrogel coating to create a versatile platform for drug delivery. To incorporate the chemotherapeutic agent, cisplatin (150 ppm) was introduced into the AgNPs-alginate mixture utilizing rapid stirring to ensure uniform distribution and encapsulation of cisplatin within the nanocomplex. A range of analytical methods, such as UV-Vis, FTIR, SEM/EDX, and Zeta potential analysis, were employed to characterize the nanocomplex. To evaluate cisplatin release kinetics from the nanocomplex, we employed an in vitro dialysis method for monitoring its sustained release. High-performance liquid chromatography (HPLC) was used for precise cisplatin release quantification, then validated by colorimetric spectrophotometry. This dual-method approach ensured accurate insights into the nanocomplex’s release dynamics, substantiating its potential in enhancing targeted cancer therapy through advanced drug delivery systems. Results The analysis through UV-Vis revealed an absorption spectrum around 410 nm, indicative of the characteristic plasmon resonance associated with silver nanoparticles. TEM provided high-resolution imagery, revealing that the size of the silver nanoparticles varied between 4 and 30 nm, averaging at 13 nm with a standard deviation of 5 nm specifically for the alginate-coated AgNPs. SEM images confirmed the anticipated spherical distribution of silver nanoparticles within the alginate hydrogel framework. EDX spectroscopy analysis further verified the incorporation of silver nanoparticles and platinum within the complex. The cisplatin release studies from this newly developed nanocomplex illustrated a prolonged, slow, and consistent release pattern, extending over days, in stark contrast to th","PeriodicalId":10690,"journal":{"name":"Clinical chemistry","volume":null,"pages":null},"PeriodicalIF":9.3,"publicationDate":"2024-10-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142369051","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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