Very Bright Marker for MHC I From BioLegend

October 05, 2023

BlueskyReddit
George Washington University

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Quality of Results

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Company:

BioLegend

Product Name:

Alexa Fluor® 647 anti-human HLA-A,B,C Antibody

Catalog Number:

311414

Clone Number:

W6/32

The goal of my research was to find increases in MHC I presentation on the cell surface of CD14+CD11c+ (Dendritic) cells when activated.

Experimental Design and Results Summary

Applications

Flow cytometry

Sample

Human dendritic cells

Primary Incubation

0.1uL Zombie Green viability stain in 100uL PBS, incubated at room temp for 15 mins

Blocking Agent

Human TruStain Fc block, 5uL diluted in 95uL FACS buffer (5% FBS in DPBS), incubated at 4C for 10 mins

Secondary Incubation

1uL of MHC I antibody diluted in 100uL of FACS buffer (5% FBS in DPBS), incubated at 4C for 20 mins

Tertiary Incubation

Cytofix buffer, 100uL for 20 mins at 4C

Detection

Flow cytometry in a BD Celesta cytometer

Results Summary

With this product, we were able to identify a shift in median fluorescence intensity of MHC I (HLA A,B,C) presented on the cell surface of dendritic cells exposed to our activating treatment. We were not able to see a distinct MHC positive or negative population, but MHC I tends to stain in a range rather than distinct populations anyway so this was expected.

DOI or PMID #

N/A

Additional Notes

N/A

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Summary

The Good

Stains very bright.

The Bad

It is very bright and likely to overlap strongly with other Alexa Fluor antibodies in your panel.

The Bottom Line

Great MHC I antibody, with a higher signal than a PE conjugate. I tried but the higher signal comes with a greater potential for overlap.

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