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Protein Size Assays on Simple Western

Simple Western Protein Size Assays add a 3rd dimension to your immunoassay

Add a 3rd Dimension to Your Immunoassay

As a capillary electrophoresis immunoassay platform, Simple Western™ provides sensitive, accurate, and reproducible quantitative immunoassay read-outs while also providing molecular weight (MW) information enabled by electrophoretic protein separation. While ELISA can only measure antibody binding as you vary experimental conditions (i.e., 2-dimensional data), Simple Western measures antibody binding activity and provides MW size information as you vary experimental conditions, adding a 3rd dimension to your immunoassay.

Size Matters Immunoassay Scientific Review

Download our Scientific Reviews Summary to learn why adding MW protein size separation to your immunoassay data matters.

Customer Success Stories

Protein Size Matters

Simple Western Protein Size Immunoassays provide antibody specificity information by monitoring off-target cross-reactivity

Specificity in Complex Lysates

Protein Assays to Guide Downstream Process Decisions

Scientists at Merck relied on Simple Western to quantify empty/full ratios of Coxsackievirus A21 (CVA21) by raising custom antibodies that target CVA21 capsid proteins. Simple Western protein size separation provided specificity of antibody reactivity to CVA21 capsid proteins. Adapted with permission (CC BY 4.0)

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Simple Western Protein Size Assay Antibody Aggregation

Isoforms, Fragments, and Aggregates

Therapeutic Antibodies

Simple Western protein size separation monitors therapeutic antibody production and aggregation directly in fed-batch and perfusion culture samples with only a single antibody. BisAb: bispecific antibody; HC: heavy chain. Adapted with permission (CC BY 4.0).2

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Simple Western Protein Size Assays Probody Cleavage Products

Cleavage Products

Probody Therapeutics

Resolving activated and intact light chain (LC) of a Probody therapeutic using the same detection antibody.3 PROBODY is a U.S.-registered trademark of CytomX Therapeutics, Inc. Adapted with permission (CC BY 4.0).3

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Simple Western Protein Size Assays Degradation Products

Degradation Products

Resolve and Quantify Full-Length Protein from Degradation Production

Immunoassay with protein size information using Simple Western is featured in a publication in Nature to measure full-length protein expression with simultaneous total protein detection (shown in blue) for accurate normalization of protein expression. Lower bands are likely degradation or incompletely synthesized products which are all detected by the same antibody and would not be resolved by ELISA or MSD assays. Adapted with permission (CC BY 4.0).4

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Simple Western Protein Size Assays Oligomer PTM

Oligomers and Post-Translational Modifications

Protein Biomarkers of Disease Pathogenesis

Teasing apart high molecular weight oligomers and deglycosylated isoforms of a biomarker implicated in multiple sclerosis, schizophrenia, and long COVID. Adapted with permission (CC BY 4.0).5

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Jess Instrument

How Simple Western Assays Can Help You

Meet Our Family of Products

Simple Western™ is a next-generation hands-free capillary immunoassay platform combining protein separation with sensitive chemiluminescence and fluorescence immunodetection using conventional primary and conjugated secondary antibodies.

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Types of Protein Size Assays


Simple Western immunoassays provide protein size separation information in addition to immunodetection of protein target(s) of interest. Simple Western merges the separative performance of capillary electrophoresis with the specificity of antibody-based detection.

Learn More about Immunoassays »


Total Protein Assays

In addition to immunoassay detection, Simple Western provides simultaneous total protein detection, which can be used to normalize protein expression data for accurate quantification of protein targets. For applications requiring normalizing protein signal, normalization to total protein is more reliable using a housekeeping protein as the normalizing denominator.

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Multiplex Assays

Get more out of your 3 µL sample! Multiplexing based on molecular weight gives you the flexibility you need to detect multiple protein targets that vary in molecular weight within the same capillary. With Jess, you have the ability to multiplex up to three channels to detect protein targets, and there is an extra channel for total protein normalization. Combine RePlex™ on Jess and Abby provides high-fidelity serial rounds of immunoprobing by removing the antibodies from the first probing cycle for a second probing cycle for immunoassay or total protein assay.

Learn More about Multiplex Assays »


Multi-Antigen Serology Assays

Size-based separation enables high-specificity multi-antigen serology testing for detailed insight into the humoral response beyond what ELISA can provide.

Learn More about Multi-Antigen Serology Assays »

Replex Diagram for Simple Western - New
  1. P Gillespie, R Rustandi, A Swartz et al. (2023) Quantitation of coxsackievirus A21 viral proteins in mixtures of empty and full capsids using capillary western Hum Gene Ther 34:66-77. PMID: 36503264.

  2. A Aziz, N Majewska et al. (2020) Perfusion reduces bispecific antibody aggregation via mitigating mitochondrial dysfunction-induced glutathione oxidation and ER stress in CHO cells Sci Rep 10:16620. PMID: 33024175.

  3. B Howng, M Winter, C LePage et al. (2021) Novel ex vivo zymography approach for assessment of protease activity in tissues with activatable antibodies Pharmaceutics 13:1390. PMID: 34575469.

  4. S Albers, E Allen, N Bharti et al. (2023) Engineered tRNAs suppress nonsense mutations in cells and in vivo Nature 618:842-848. PMID: 37258671.

  5. B Charvet, J Pierquin, J Brunel et al. (2021) Human endogenous retrovirus type W envelope from multiple sclerosis demyelinating lesions shows unique solubility and antigenic characteristics Virol Sin 36:1006-1026. PMID: 33770381.