Western Blot (Protein Immunoblot)
Definition
SDS-polyacrylamide gel electrophoresis (SDS-PAGE) of proteins followed by electrotransfer to a PVDF or nitrocellulose membrane, then antibody-based detection of a specific protein at its molecular weight position. Used forensically for PSA confirmation, species identification, and prion-protein characterisation.
- First step
- SDS-PAGE separation of proteins
- Second step
- Electrotransfer to PVDF or nitrocellulose membrane
- Detection
- Antibody binding at the protein's molecular weight
- Forensic uses
- PSA confirmation, species ID, prion characterisation
Common questions
How does Western blot differ from Southern and Northern blot?+
Southern blot detects DNA and Northern blot detects RNA using nucleic acid hybridisation, while Western blot detects protein using antibody binding after gel separation and membrane transfer; the target molecule and detection chemistry differ across all three.
Why is Western blot used to confirm PSA in sexual assault casework rather than a simpler test?+
A confirmatory Western blot verifies that a positive screening result truly reflects the target protein at its expected molecular weight, reducing the risk of a false positive from a cross-reacting substance in a presumptive test.
Related terms
- Microarray (Gene Chip / SNP Chip)
- A solid-phase hybridisation platform carrying thousands to millions of oligonucleotide probes at spatially registered positions. Hybridisation of a labelled target simultaneously queries...
- Minisatellite (VNTR)
- A hypervariable genomic locus characterised by tandem repeats of a 10-60 bp core sequence, with the number of repeat units varying between...
- Molecular Hybridisation
- The formation of a double-stranded nucleic acid molecule from two complementary single strands, driven by hydrogen-bond formation between base pairs (A-T, G-C...
- mRNA Body-Fluid Profiling
- The RT-PCR-based descendant of Northern blotting: tissue-specific messenger RNA transcripts are reverse-transcribed and amplified from crime-scene stains to identify body fluid of...
- Southern Blot
- A technique (E.M. Southern, 1975) in which restriction-digested DNA fragments are separated by gel electrophoresis, transferred to a membrane, and hybridised with...
- Stringency
- The extent to which hybridisation conditions favour only perfectly complementary probe-target pairs over partially mismatched ones. High stringency (elevated temperature and/or low...