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Western Blotting

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.

Southern Blot

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.
Denatured DNA fragments must be transferred onto a carrier membrane from the gel to make it accessible to a probe - a small ssDNA fragment complementary to the target DNA...

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Related Experiment Video

Updated: Nov 30, 2025

Primer-Free Aptamer Selection Using A Random DNA Library

Primer-Free Aptamer Selection Using A Random DNA Library

Published on: July 26, 2010

Aptamer-Based Western Blot for Selective Protein Recognition.

Yao Wang^1,2,3, Zhe Li^1,4, Hanyang Yu^1,2,3

¹Department of Biomedical Engineering, College of Engineering and Applied Sciences, Jiangsu Key Laboratory of Artificial Functional Materials, Nanjing University, Nanjing, China.

Frontiers in Chemistry

|November 16, 2020

Summary

This summary is machine-generated.

DNA aptamers offer a cost-effective alternative to antibodies for Western blotting. These aptamers selectively label target proteins like GST, MBP, and His-tag, improving efficiency and reducing background noise in protein analysis.

Keywords:

Western blot aptamer functional nucleic acid in vitro selection protein recognition

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Published on: October 6, 2022

Area of Science:

Molecular Biology
Biotechnology
Biochemistry

Background:

Selective protein recognition is crucial for molecular biology techniques like Western blotting and ELISA.
Antibodies, commonly used for protein detection, are expensive and require long incubation times.
Aptamers offer a potentially more affordable and faster alternative to antibodies.

Purpose of the Study:

To isolate and characterize DNA aptamers for selective recognition of common tag proteins (GST, MBP, His-tag).
To evaluate the efficacy of aptamer-based Western blotting compared to traditional antibody-based methods.
To demonstrate the broad applicability of these aptamers in protein analysis.

Main Methods:

In vitro selection was employed to isolate DNA aptamers against GST, MBP, and His-tag.
Aptamer binding affinities were determined.
Aptamer-based Western blotting was performed and compared to antibody-based methods.
The aptamers' ability to recognize fusion proteins with the same tags was assessed.

Main Results:

Novel DNA aptamers (G1, M1, H1) were generated, exhibiting specific binding to GST, MBP, and His-tag, respectively, with nanomolar affinities.
Aptamer-based Western blotting resulted in a cleaner background and superior selectivity compared to antibody-based methods.
The identified aptamers successfully recognized various fusion proteins containing the target tags.

Conclusions:

DNA aptamers are effective molecular tools for selective protein recognition in Western blotting.
Aptamer-based approaches offer a time- and cost-effective alternative to conventional antibody-based techniques.
The developed aptamers have broad potential applications in detecting tagged proteins and fusion proteins.