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Related Concept Videos

Western Blotting01:15

Western Blotting

Western blotting is an analytical technique for protein identification. It has various applications in immunology and medicine, including detecting diseases like bovine spongiform encephalopathy, mad cow disease, and human and feline immunodeficiency virus from biological samples.
The technique begins with separating proteins from the sample using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), followed by protein transfer, immunoblotting, and finally, protein detection.
Immunogold Electron Microscopy01:20

Immunogold Electron Microscopy

Immunoelectron microscopy utilizes immunogold labeling of endogenous proteins with specific antibodies to detect and localize these proteins in cells and tissues. The procedure provides insights into the distribution and quantification of protein under different stimulation conditions offering clues about their functions. Conjugating highly electron-dense gold particles with primary or secondary antibodies allow antigen detection on and within cells, with high resolution and specificity.
Labeling DNA Probes03:31

Labeling DNA Probes

DNA probes are fragments of DNA labeled with a reporter tag to enable their detection or purification. The resulting labeled DNA probes can then hybridize to target nucleic acid sequences through complementary base-pairing, and may be used to recover or identify these regions.
Radioisotopes, fluorophores, or small molecule binding partners like biotin or digoxigenin, are the most widely used reporter tags for labeling DNA probes. These labels can be attached to the probe DNA molecule via...
Enzyme-Linked Immunosorbent Assay01:33

Enzyme-Linked Immunosorbent Assay

In 1971, Peter Perlman and Eva Engvall developed an Enzyme-linked immunosorbent assay (ELISA or EIA). ELISA differs from western blot in that the assays are conducted in microtiter plates or in vivo rather than on an absorbent membrane.
There are many different types of ELISAs, but they all involve an antibody molecule whose constant region binds an enzyme, leaving the variable region free to bind its specific antigen.  Enzyme-substrate reaction allows the antigen to be visualized or quantified.
Southern Blot02:57

Southern Blot

Agarose gel electrophoresis is very useful in separating DNA fragments by size. Running a DNA ladder containing fragments of the known length alongside the sample helps determine the approximate length of the sample DNA fragments. However, additional steps are needed to verify the sequence identity of the sample DNA fragments.
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Related Experiment Video

Updated: Jun 23, 2026

Immunoblot Analysis
16:01

Immunoblot Analysis

Published on: June 20, 2008

Immunoblotting using radiolabeled reagents for detection.

Holger Bartsch1, Claudia Franke, Michael Bachmann

  • 1Institute Immunology, MTZ, Carl Gustav Carus University TU Dresden, Medical Faculty, Fiedlerstrasse 42, D-01307, Dresden, Germany.

Methods in Molecular Biology (Clifton, N.J.)
|April 21, 2009
PubMed
Summary
This summary is machine-generated.

Radioiodinated antibodies offer a rapid, sensitive alternative to enzyme-labeled antibodies for immunoblots. This method enables quantitative evaluation with low background during autoantigen purification.

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Area of Science:

  • Biochemistry
  • Immunology
  • Analytical Chemistry

Background:

  • Traditional immunoblots utilize enzyme-labeled antibodies, converting soluble substrates to insoluble products.
  • This process can be time-consuming and may result in high background noise.
  • Quantitative evaluation can also be challenging with conventional methods.

Purpose of the Study:

  • To present a simple, rapid, and sensitive alternative for immunodetection.
  • To introduce the use of radiolabeled antibodies for quantitative evaluation.
  • To demonstrate the application of iodinated secondary antibodies in autoantigen purification via HPLC.

Main Methods:

  • Development of immunoblots using radiolabeled antibodies (specifically, iodinated secondary antibodies).
  • Application of the method for immunodetection of an autoantigen.
  • Utilizing High-Performance Liquid Chromatography (HPLC) for purification and detection.

Main Results:

  • The radiolabeling method provides a sensitive alternative to enzyme-based detection.
  • Low background signals were achieved, facilitating clearer results.
  • The method allows for rapid quantitative evaluation of autoantigens.
  • Successful application in autoantigen purification during HPLC.

Conclusions:

  • Radiolabeled antibodies, particularly iodinated secondary antibodies, offer a viable and advantageous alternative for immunoblotting.
  • This technique enhances sensitivity, reduces background, and enables rapid quantitative analysis.
  • The described method is effective for autoantigen detection and purification using HPLC.