ReviewAmerican journal of physiology. Cell physiology2015
Considerations when quantitating protein abundance by immunoblot.
Review in American journal of physiology. Cell physiology, 2015. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 65 papers, 1 of them a synthesis that pooled it.
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Who cites it
65 citing papers in PubMed, 1 synthesis or guideline pooled it, 104 citations in OpenAlex.
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- Kidney-specific WNK1 amplifies kidney tubule responsiveness to potassium via WNK body condensates.The Journal of clinical investigation · 2025Article
- Titration-WB: A methodology for accurate quantitative protein determination overcoming reproducibility errors.PloS one · 2025Article
- Optimizing renal transporter immunodetection: consequences of freeze-thaw during sample preparation.American journal of physiology. Renal physiology · 2024Article
- Improving rigor and reproducibility in western blot experiments with the blotRig analysis.Scientific reports · 2024Article
- Recommendations for Development and Validation of a Fit-For-Purpose Biomarker Assays Using Western Blotting; An-AAPS Sponsored Initiative to Harmonize Industry Practices.The AAPS journal · 2024Article
- Evolution of ion transporter NaiScience · 2024Article
- Detection and quantification of C-terminally tagged proteins by in-gel fluorescence.Scientific reports · 2024Article
- Guidelines on antibody use in physiology research.American journal of physiology. Renal physiology · 2024Article
- Data-Independent Acquisition Proteomics and N-Terminomics Methods Reveal Alterations in Mitochondrial Function and Metabolism in Ischemic-Reperfused Hearts.Journal of proteome research · 2024Article
- Monitoring phosphorylation and acetylation of CRISPR-mediated HiBiT-tagged endogenous proteins.Scientific reports · 2024Article
- Dysregulation of the WNK4-SPAK/OSR1 pathway has a minor effect on baseline NKCC2 phosphorylation.American journal of physiology. Renal physiology · 2024Article
- Chemokines and lymphocyte homing in Sjögren's syndrome.Frontiers in immunology · 2024Review
- Comparing ATPase activity of ATP-binding cassette subfamily C member 4, lamprey CFTR, and human CFTR using an antimony-phosphomolybdate assay.Frontiers in pharmacology · 2024Article
5 more citing papers are in PubMed but not listed here.
Corrections and comments
PubMed lists nothing against this paper. Absence here is not a guarantee, only a check that was made.
Authors and funding
4 authors at 2 institutions in 1 country.
Funding
Abstract
The development of the immunoblot to detect and characterize a protein with an antisera, even in a crude mixture, was a breakthrough with wide-ranging and unpredictable applications across physiology and medicine. Initially, this technique was viewed as a tool for qualitative, not quantitative, analyses of proteins because of the high number of variables between sample preparation and detection with antibodies. Nonetheless, as the immunoblot method was streamlined and improved, investigators pushed it to quantitate protein abundance in unpurified samples as a function of treatment, genotype, or pathology. This short review, geared at investigators, reviewers, and critical readers, presents a set of issues that are of critical importance for quantitative analysis of protein abundance: 1) Consider whether tissue samples are of equivalent integrity and assess how handling between collection and assay influences the apparent relative abundance. 2) Establish the specificity of the antiserum for the protein of interest by providing clear images, molecular weight markers, positive and negative controls, and vendor details. 3) Provide convincing evidence for linearity of the detection system by assessing signal density as a function of sample loaded. 4) Recognize that loading control proteins are rarely in the same linear range of detection as the protein of interest; consider protein staining of the gel or blot. In summary, with careful attention to sample integrity, antibody specificity, linearity of the detection system, and acceptable loading controls, investigators can implement quantitative immunoblots to convincingly assess protein abundance in their samples.
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Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.