Protein expression and purification最新文献

{"title":"Bioinformatics analysis and preparation of polyclonal antibodies against the extracellular region of the Langya Virus attachment protein","authors":"Shuxing Qiu ,&nbsp;Xing Yin ,&nbsp;Jinjiao He ,&nbsp;Jiayou Zhang ,&nbsp;Jinjing Guo ,&nbsp;Yonghua Qi ,&nbsp;Feifan Liu","doi":"10.1016/j.pep.2026.106888","DOIUrl":"10.1016/j.pep.2026.106888","url":null,"abstract":"<div><div>The Langya virus attachment glycoprotein (LayV-G) is a major surface glycoprotein critical for host cell entry, membrane fusion, and the induction of neutralizing antibody production. A highly specific, reliable, and cost-effective anti-LayV-G antibody is urgently needed to comprehensively assess its multifunctional role in the viral life cycle. Here, we report the first successful production of LayV-G protein and corresponding polyclonal antibodies (pAbs) using a prokaryotic expression system. In this study, the extracellular domain of LayV-G containing strong antigenic epitopes was predicted using bioinformatics approaches and selected for recombinant antigen production, followed by codon optimization and synthesis of the candidate protein-encoding gene. We then constructed the truncated attachment glycoprotein (tG) with a C-terminal histidine tag (His-tag), (pET-22b-tG/his) recombinant plasmid and expressed the protein in <em>E. coli</em> BL21 (DE3) competent cells. SDS–PAGE and subsequent tandem mass spectrometry revealed the molecular weight of the tG/his recombinant protein to be ∼64 kDa (kD). Subsequently, the targeted protein was used to immunize New Zealand white rabbits and generate pAbs. The specificity and titer of the pAbs were determined by Western blotting, indirect fluorescence assay (IFA), and indirect ELISA. The antibody exhibited high specificity for tG/his in prokaryotic cells (Western blotting), and LayV-G expressed by the recombinant baculovirus system (IFA). In conclusion, the LayV-G protein and pAbs targeting it lays the foundation for further investigations into LayV-G structure and function, pathogenic LayV molecular mechanisms, and rapid LayV detection methods.</div></div>","PeriodicalId":20757,"journal":{"name":"Protein expression and purification","volume":"240 ","pages":"Article 106888"},"PeriodicalIF":1.2,"publicationDate":"2026-01-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"146044040","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Scalable production and characterization of recombinant pneumococcal histidine triad protein D in Escherichia coli for vaccine development","authors":"Prashant Jadhav ,&nbsp;Hitesh Malviya ,&nbsp;Jyoti Gokhale","doi":"10.1016/j.pep.2026.106889","DOIUrl":"10.1016/j.pep.2026.106889","url":null,"abstract":"<div><div><em>Streptococcus pneumoniae</em> causes 1.5 million deaths annually, primarily in low-income countries. Pneumococcal histidine triad protein D (PhtD), a conserved 92–110 kDa surface protein, is a promising serotype-independent vaccine candidate due to its roles in adherence, zinc homeostasis, and complement evasion. This study optimized scalable production of His-tagged PhtD in <em>Escherichia coli</em> BL21(DE3). The PhtD gene (serotype 2, strain D39) was cloned into pET30a(+), and expression was optimized using a full factorial design of experiments, achieving up to 19.8 mg/L soluble PhtD (&gt;90 % purity) with 0.1 mM IPTG at OD₅₉₀ = 0.2 for 2 h. Lysozyme treatment (0.3 mg/mL) increased yield to 22 mg/L, a 120 % improvement over the control (10 mg/L). Purification involved sonication, ultrafiltration, and IMAC (80–100 mM imidazole wash, 200–300 mM elution), with 1 mM PMSF preventing ∼30 % yield loss from proteolysis. PhtD's identity (105–110 kDa), secondary structure (15.5 % helix, 30.4 % sheet), and thermal stability (Tm ∼102 °C) were confirmed by SDS-PAGE, Western blotting, FTIR, circular dichroism, NMR, and DSC. This optimized, statistically guided process delivers the highest reported yield of soluble full-length PhtD in cytoplasmic <em>E. coli</em> with documented native-like folding and exceptional stability (Tm = 102 °C), providing a robust and scalable platform for serotype-independent pneumococcal vaccine development.</div></div>","PeriodicalId":20757,"journal":{"name":"Protein expression and purification","volume":"240 ","pages":"Article 106889"},"PeriodicalIF":1.2,"publicationDate":"2026-01-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145981457","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Additive-based protein stabilization for stress-prone and unstable proteins","authors":"Tomoto Ura,&nbsp;Takumi Nakamura,&nbsp;Moe Iijima,&nbsp;Toya Yoshida,&nbsp;Kentaro Shiraki","doi":"10.1016/j.pep.2026.106885","DOIUrl":"10.1016/j.pep.2026.106885","url":null,"abstract":"<div><div>Protein-stabilizing additives have traditionally been investigated under dilute solution conditions using simple monomeric proteins as model systems. While such studies have clarified fundamental additive–protein interactions, their direct applicability to practical research and industrial contexts remains limited. This review leverages insights from model systems to summarize additive-based stabilization strategies in two major directions. First, we address process-related stresses, focusing on high-concentration conditions and interfacial stress frequently encountered during protein handling. Second, we highlight strategies for protein classes that are challenging to stabilize or purify, including membrane proteins, viral spike proteins, and intrinsically disordered proteins. This review aims to encourage additive use when proteins are difficult to stabilize under challenging conditions.</div></div>","PeriodicalId":20757,"journal":{"name":"Protein expression and purification","volume":"240 ","pages":"Article 106885"},"PeriodicalIF":1.2,"publicationDate":"2026-01-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145960075","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Development and optimization of the hypoglycemic activity of an α-glucosidase inhibitory protein from a UV-induced mutant strain of Streptomyces griseorubens","authors":"Mohammed Alosaimi ,&nbsp;Waleed Abdulkhair ,&nbsp;Jamal Asseri ,&nbsp;Abdulmajeed Almuqrin ,&nbsp;Fatemah AlMalki","doi":"10.1016/j.pep.2026.106878","DOIUrl":"10.1016/j.pep.2026.106878","url":null,"abstract":"<div><div>This study investigated the isolation, enhancement, and optimization of an α-glucosidase inhibitory protein with hypoglycaemic activity from <em>Streptomyces</em> species obtained from tomato-cultivated soils rich in actinomycetes. A total of 110 <em>Streptomyces</em> isolates were recovered from ten soil samples collected in Sakha City. Sixteen isolates exhibited notable α-glucosidase inhibitory activity, with isolate Strep-5 showing the highest potency. Morphological, biochemical, physiological, and molecular analyses identified Strep-5 as <em>Streptomyces griseorubens</em>. Scanning electron microscopy revealed spiral spore chains with smooth, ellipsoidal spores, and 16S rRNA phylogenetic analysis confirmed a close relationship to <em>S. griseorubens</em> (≥90 % bootstrap support). UV mutagenesis markedly enhanced the inhibitory activity, with a 20-s exposure producing the most active mutant. Optimal production occurred in yeast–malt extract broth at 35 °C, pH 7.5, and 160 rpm after 72 h of incubation. Sequential purification using ammonium sulfate precipitation, ion-exchange, and gel filtration chromatography yielded an 8.5-fold purification and 40.8 % recovery. The purified enzyme displayed a single 70 kDa band on SDS–PAGE, confirming homogeneity, and exhibited total and specific activities of 21,000 U mL⁻¹ and 1020 U mg⁻¹, respectively. UV mutagenesis significantly improved the α-glucosidase inhibitory activity of <em>S. griseorubens</em> Strep-5, producing a stable, high-molecular-weight protein with strong antidiabetic potential. This work demonstrates the successful isolation, mutagenic enhancement, and optimization of a microbial α-glucosidase inhibitor from <em>S. griseorubens</em>. The findings highlight its potential as a promising biotechnological and therapeutic agent for diabetes management.</div></div>","PeriodicalId":20757,"journal":{"name":"Protein expression and purification","volume":"240 ","pages":"Article 106878"},"PeriodicalIF":1.2,"publicationDate":"2026-01-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145945919","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Expression and production of recombinant SARS-CoV-2 RBD protein in E. coli system: comparison of performance, functionality and stability in three strains","authors":"Romina Golafshan ,&nbsp;Mahmoudreza Aghamali ,&nbsp;Mohammad Javad Rasaee","doi":"10.1016/j.pep.2025.106877","DOIUrl":"10.1016/j.pep.2025.106877","url":null,"abstract":"<div><div>The worldwide COVID-19 pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-Cov-2). The receptor binding domain (RBD) of SARS-Cov-2 is located on the spike protein, which binds and interacts with ACE2. The RBD serve as a crucial target for the development of virus-neutralizing antibodies, vaccinations, and inhibiting agents. The goal of this study was to synthesis recombinant RBD (rRBD) protein and evaluate its expression, function, and stability in different strains of <em>Escherichia coli</em> (<em>E. coli</em>). The rRBD domain gene sequence was engineered utilizing in silico methodologies. The optimized sequence was inserted into pET-28a vector using <em>Xho</em>I and <em>Eco</em>RI restriction sites. Three <em>E. coli</em> expression strains BL21 (DE3), Rosetta-gami, and Shuffle T7 were chosen for protein synthesis. Cloning of the target gene fragment was confirmed using PCR. The validated recombinant vector was cloned into three <em>E. coli</em> strains. SDS-PAGE and Western blot analysis were used to examine the expression of the rRBD. Far-UV circular dichroism (CD) spectroscopy was used to examine secondary structure. ELISA experiments quantified the interaction of rRBD protein with HRP-conjugated anti-His antibody and serum from COVID-19 patients detected with anti-human immunoglobulin-HRP labeled. Protein stability was assessed under different temperature conditions. The rRBD protein was best expressed in all three bacterial strains at 1 mM concentration of IPTG. A 14 kDa band corresponding to the rRBD protein was identified in all three bacterial strains via Western blot. BL21 had the greatest amount of rRBD expression. The rRBD generated from Rosetta-gami exhibited the most robust affinity for anti-His antibodies. All three expressed proteins exhibited similar secondary structure, characterized by a predominance of random coil and β-sheet content. They also exhibited similar affinity to antibodies against SARS-Cov-2 generated from patients. Storage at −20 °C and 4 °C with suitable glycerol concentrations ensured optimal long-term protein stability. This study reveals that <em>E. coli</em> is still a good and flexible host system to produce functional SARS-Cov-2 RBD protein. Even though the strains had different levels of expression and binding efficacy, all of the proteins they made had comparable structures and antigenic properties. The findings validate the feasibility of using bacterial expression systems for the cost-effective production of RBD-based diagnostic reagents.</div></div>","PeriodicalId":20757,"journal":{"name":"Protein expression and purification","volume":"239 ","pages":"Article 106877"},"PeriodicalIF":1.2,"publicationDate":"2026-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145896848","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Detection of human PD-1 on T-cells using a fusion protein consisting of an anti-PD-1 single chain variable fragment linked to superfolder GFP","authors":"Jong Shin ,&nbsp;David Sanford ,&nbsp;Alex A. Bohm ,&nbsp;William Bachovchin ,&nbsp;Peter A. Bullock","doi":"10.1016/j.pep.2025.106876","DOIUrl":"10.1016/j.pep.2025.106876","url":null,"abstract":"<div><div>Monoclonal antibodies that disrupt the interaction between T-cell derived PD-1 and immunosuppressive ligands, such as PD-L1, have dramatically improved approaches to cancer therapy. One such antibody is termed Nivolumab (trade name Opdivo) that selectively targets PD-1. We previously described a Nivolumab-based anti-PD-1 single chain variable fragment (scFv), termed the anti-PD-1 double mutant (dm) scFv, that tightly binds to PD-1 and blocks the interaction between PD-1 expressed on T-cells and PD-L1 on CHO cells. Extending these experiments, we now report a fluorescent reporter formed by the fusion of the anti-PD-1dm with superfolder (sf) GFP. Immunofluorescence and flow cytometry studies established that the anti-PD-1dm-sfGFP fusion protein can be used to detect PD-1 on the surface of PD-1 expressing Jurkat cells. Furthermore, using a humanized mouse strain that expresses human PD-1 and PD-L1, we report that when combined with an anti-CD3 antibody, the anti-PD-1dm-sfGFP can be used to identify PD-1 expressing T-cells within mouse tumor biopsies. Thus, we have developed a reagent for the quantitative determination of activated T-cells in tumor biopsies. Finally, we report that an additional fluorescent probe for the detection of PD-1 <em><u>in vivo</u></em> can be generated by linking, via maleimide chemistry, a derivative of the anti-PD-1dm to the near-infrared dye IR800.</div></div>","PeriodicalId":20757,"journal":{"name":"Protein expression and purification","volume":"240 ","pages":"Article 106876"},"PeriodicalIF":1.2,"publicationDate":"2025-12-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145865191","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Novel anti-NS1 human single domain antibody from dengue infected NS1 positive patients","authors":"Odedara Mitalben Bhikhabhai,&nbsp;Hemant Kumar","doi":"10.1016/j.pep.2025.106875","DOIUrl":"10.1016/j.pep.2025.106875","url":null,"abstract":"<div><div>Viruses of the flavivirus family significantly affect human health worldwide. Dengue virus (DENV), a member of the flavivirus family, causes pathogenic symptoms like high fever, severe headache and body pain upon infection. In some cases, it develops into severe dengue that is fatal. As a crucial pillar of immune response antibodies are produced against different epitopes of the dengue proteins and a collective response is generated by targeting critical proteins. A vaccine against dengue is still to be developed and new antibodies are the need of the hour. In our work we have isolated novel full length heavy chain variable region genes from peripheral blood mononuclear cell (PMBC) of NS1 positive DENV infected patients. The sequencing suggests that the isolated heavy chains to be novel and not yet reported. The sequence verified clones were sub cloned into expression vector pET-28a(+) for protein expression in <em>E. coli</em>. Selected heavy chains were overexpressed and purified to near homogeneity and further specific binding of purified heavy chain variable region with rNS1 was confirmed through competitive inhibition ELISA, dose dependent competitive inhibition ELISA and western blotting. ITC further confirmed the binding of rNS1 and SdAb3 with favourable free energy (ΔG = −9.22 kcal/mol). Identity of purified protein was confirmed by western blotting through anti-His antibody. It was found that a large amount of the purified protein was going in insoluble fraction and a protein refolding strategy was optimized with different buffer conditions. A buffer containing arginine, histidine and sucrose was found to be most efficient in solubilization of protein from insoluble fraction.</div></div>","PeriodicalId":20757,"journal":{"name":"Protein expression and purification","volume":"239 ","pages":"Article 106875"},"PeriodicalIF":1.2,"publicationDate":"2025-12-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145850662","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Expression of recombinant preenzyme Blo t 1 allergen in Escherichia coli and its IgE reactivity","authors":"Chi Linh Thi Nguyen ,&nbsp;Uyen Quynh Nguyen ,&nbsp;Phuong Mai Vu ,&nbsp;Truong Huu Nguyen ,&nbsp;Phuong Hoang Nguyen ,&nbsp;Chi Quynh Thi Do ,&nbsp;Vinh Van Hoang","doi":"10.1016/j.pep.2025.106872","DOIUrl":"10.1016/j.pep.2025.106872","url":null,"abstract":"<div><div><em>Blomia tropicalis</em> is a predominant house dust mite in tropical-subtropical climates, and its allergen Blo t 1 is recognized as a key trigger of allergic responses. This study aimed to express, purify recombinant Blo t 1 from <em>B. tropicalis</em> collected in Vietnam and assess its IgE-binding capacity using Vietnamese allergic sera. The cDNA encoding Blo t 1 was amplified from local samples, cloned into pET32b (+), and expressed in <em>Escherichia coli</em> BL21 (DE3). IgE-binding properties were evaluated through dot blot and Western blot assays. The recombinant Blo t 1 showed anomalous SDS-PAGE migration at ∼63 kDa, comprising ∼38.11 kDa from 333 amino acids of the pre-enzyme form of Blo t 1 and ∼19.47 kDa derived from vector-associated sequences, including His-tag. The protein was successfully expressed and purified using His-tag affinity chromatography. Dot blot analysis showed that 19/30 sera (63.33 %) from <em>B. tropicalis</em>-sensitized patients reacted with the recombinant protein, whereas none of the 24 control sera did. Western blot confirmed IgE-binding specificity. The recombinant Blo t 1 was purified under denaturing conditions, so the native three-dimensional structure is unlikely to be retained, and the observed IgE reactivity mainly reflects linear epitope recognition. Even so, the protein represents a useful research material corresponding to a Blo t 1 variant circulating in Vietnam. Along with expanding the limited molecular and IgE-reactivity data available for <em>B. tropicalis</em> in Southeast Asia, it also provides a basis for studies on allergen structure, epitope features, and early exploratory work related to allergen-specific immunotherapy and SIT-oriented approaches.</div></div>","PeriodicalId":20757,"journal":{"name":"Protein expression and purification","volume":"239 ","pages":"Article 106872"},"PeriodicalIF":1.2,"publicationDate":"2025-12-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145828358","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Expression and functional characterization of recombinant soluble CD40 Ligand in prokaryotic systems","authors":"Qi Xu ,&nbsp;Xinying He ,&nbsp;Jiale Bai ,&nbsp;Hanlin Liu ,&nbsp;Tao Hong ,&nbsp;Suying Dang ,&nbsp;Wei Zhang","doi":"10.1016/j.pep.2025.106874","DOIUrl":"10.1016/j.pep.2025.106874","url":null,"abstract":"<div><h3>Objective</h3><div>To develop an optimized prokaryotic expression system for producing functional soluble CD40 ligand (sCD40L) and characterize its hemostatic activity.</div></div><div><h3>Methods</h3><div>The codon-optimized <em>sCD40L</em> gene (encoding residues 113–261) was synthesized and cloned into pET28a vector via BamHI/XhoI sites. After transformation into <em>E. coli</em> BL21(DE3), soluble expression was induced with 0.5 mM IPTG at 20 °C for 12 h. The recombinant protein was purified using nickel-affinity chromatography and analyzed by SDS-PAGE and Western blot. Biological activity was assessed through in vitro platelet aggregation and in vivo hemostasis assays.</div></div><div><h3>Results</h3><div>The pET28a-sCD40L expression vector was successfully constructed, and the protein purity reached more than 90 %. Western blot confirmed the expected ∼20 kDa sCD40L. Functionally, the recombinant sCD40L enhanced platelet aggregation in a dose-dependent manner, and significantly reduced secondary bleeding time in mice.</div></div><div><h3>Conclusion</h3><div>Our optimized prokaryotic system efficiently produces functional sCD40L without requiring eukaryotic post-translational modifications, providing a valuable tool for studying CD40L-mediated hemostasis and developing potential therapeutic applications.</div></div>","PeriodicalId":20757,"journal":{"name":"Protein expression and purification","volume":"239 ","pages":"Article 106874"},"PeriodicalIF":1.2,"publicationDate":"2025-12-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145786906","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"The superiority of MabSelect VL, Cytiva's second-generation Protein L resin, over three other affinity resins in purifying a VHH-containing trispecific antibody","authors":"Ruomei Lv ,&nbsp;Wenjing Qi ,&nbsp;Yifeng Li","doi":"10.1016/j.pep.2025.106873","DOIUrl":"10.1016/j.pep.2025.106873","url":null,"abstract":"<div><div>Affinity chromatography is widely utilized for initial product capture in antibody purification. Besides Protein A resins, other affinity resins that bind different subdomains of an antibody have also been developed. For example, Protein L resin, KappaSelect, LambdaFabSelect and Praesto 70 CH1 selectively bind the variable region of kappa light chain (LC), constant region of kappa LC, constant region of lambda LC and CH1 domain of the heavy chain (HC), respectively. Affinity media with distinct specificities provide more options for antibody purification. For a given bispecific antibody (bsAb) purification case, depending on the composition difference between the product and byproducts, one affinity resin may outperform the others for byproduct removal. In the current study, we compared four affinity resins, Praesto Jetted A50 (a Protein A resin), Praesto 70 CH1, KappaSelect and MabSelect VL (Cytiva's second-generation Protein L resin), for their effectiveness in removing byproducts associated with a VHH-containing trispecific antibody (TsAb). The data suggested that the three non-Protein A resins outperformed the Protein A resin for byproduct removal. Among the three non-Protein A resins, MabSelect VL provided the best byproduct clearance, and it removed half-antibody, homodimers and most aggregates. This work reconfirms the necessity of screening different affinity resins with distinct specificities for achieving the best separation.</div></div>","PeriodicalId":20757,"journal":{"name":"Protein expression and purification","volume":"239 ","pages":"Article 106873"},"PeriodicalIF":1.2,"publicationDate":"2025-12-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145794505","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}