Antibody Labeling With Alexa Fluorophore (AF660) Conjugation Kit

March 29, 2018

University of Rochester Medical Center
Microbiology & Immunology
Research Scientist

Overall

Quality of Results

Ease-of-Optimization

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Company:

Thermo Fisher Scientific

Product Name:

Alexa Fluor 660 Protein Labeling Kit

Catalog Number:

A20171

Alexa fluorophore-conjugated antibody is popular in flow cytometry and immunofluorescence microscopy. However, antibodies conjugated with a certain specific alexa fluorphore species are not commercially available. In this case, you are required to label the antibody yourself. Here, I report a review of labeling of an antibody with Alexa fluorophore 660 using a kit purchased from Invitrogen.

Experimental Design and Results Summary

Application

Fluorophore labeling of an antibody to use it for flow cytometry

Starting Material

Protein/antibody

Protocol Overview

I labeled anti-Ly6G (rat monoclonal IgG) antibody with Alexa fluorophore 660 using a kit from Invitrogen. The kit allows three trials of labeling of mg level of a protein/antibody. The kit includes everything required for the labeling, so you only need unlabeled target protein/antibody. The procedure is very easy to follow to everybody who has no experience and knowlege of protein chemistry.

Tips

This series of labeling kit is for whom needs to label his/her home-made protein/antibody or you don't find an antibody conjugated with the fluorophore that you want.

Results Summary

Unlabeled Ly6G antibody from BioXcell was conjugated with Alexa fluorophore 660 following the instruction included. I could obtain approximately 1 mg of brightly labeled antibody which was ready to use within a couple of hours.

DOI or PMID #

N/A

Additional Notes

N/A

Image Gallery

Summary

The Good

Easy to follow; the conjugation is highly efficient; labeled antibody was fully functional in flow cytometry

The Bad

You may need to change the buffer such as using dialysis

The Bottom Line

Basically, this kit allows anybody to easily label his/her protein/antibody.

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