Journal article
Stain-Free total-protein normalization enhances the reproducibility of Western blot data
Analytical biochemistry, Vol.654, p.114840
10/01/2022
DOI: 10.1016/j.ab.2022.114840
PMID: 35931182
Abstract
We compared the accuracy of three common methods of total protein normalization. The Stain-Free method was accurate across different types/brands of western blotting membrane and for various protein loads, unlike Ponceau S and Amido Black. Normalizing to the housekeeping proteins Actin and β-Tubulin could match the accuracy of the Stain-Free method. However, compared to Actin or β-Tubulin, normalizing to the Stain-Free signal reduced variability that led to enhanced reproducibility and a reduction in the number of samples needed to obtain statistically significant results by >50%. Stain-Free normalization can enhance the reproducibility and hence the confidence in Western Blot data.
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•Total-protein normalization methods vary in accuracy by type/brand of immunoblotting membrane and amount of protein load.•Housekeeping protein normalization can match the accuracy of Stain-Free total protein normalization but with less precision.•Stain-Free total protein normalization can enhance the reproducibility and hence the confidence in Western Blot data.
Details
- Title: Subtitle
- Stain-Free total-protein normalization enhances the reproducibility of Western blot data
- Creators
- Abigail Maloy - University of PittsburghShaun Alexander - University of PittsburghAlexis Andreas - University of PittsburghToru Nyunoya - University of PittsburghDivay Chandra - University of Pittsburgh
- Resource Type
- Journal article
- Publication Details
- Analytical biochemistry, Vol.654, p.114840
- DOI
- 10.1016/j.ab.2022.114840
- PMID
- 35931182
- NLM abbreviation
- Anal Biochem
- ISSN
- 0003-2697
- eISSN
- 1096-0309
- Publisher
- Elsevier Inc
- Language
- English
- Date published
- 10/01/2022
- Academic Unit
- Internal Medicine
- Record Identifier
- 9985214169702771
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