Great Donkey anti-Sheep Alexa 555 Secondary Antibody for IHC

April 19, 2016

Center for Hearing & Deafness
University at Buffalo
Principal Investigator

Overall

Quality of Results

Ease-of-Optimization

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Company:

Invitrogen

Product Name:

Donkey anti-Sheep IgG (H+L) Secondary Antibody

Catalog Number:

A21436

We needed a good secondary antibody for double immunostaining on mouse xenografts. We already had a rabbit primary antibody and a mouse antibody would lead to mouse-on-mouse background problems. So, we chose a sheep primary antibody and paired it with this Donkey anti-Sheep Alexa 555 secondary antibody.

Experimental Design and Results Summary

Applications

Immunohistochemistry

Sample

Paraffin section of a mouse xenograft

Primary Incubation

Sheep anti-Estrogen Receptor, (R&D Systems AF5715) incubated at 2 ug/ml for one hour at room temperature.

Blocking Agent

NA

Secondary Incubation

Life Technologies A21436 Donkey anti-Sheep Alexa 555 was used at 2 ug/ml for 45 minutes at room temperature.

Tertiary Incubation

Vectashield mounting medium with DAPI (blue fluorescent nuclei)

Detection

Fluorescence microscopy using red and blue filters

Results Summary

The staining was very specific, with good contrast and low background. We could easily detect ER+ tumor cells in the xenograft.

Additional Notes

None

Image Gallery

Summary

The Good

Easy to optimize

The Bad

None

The Bottom Line

This Donkey anti-Sheep IgG (H+L) Secondary Antibody, Alexa Fluor® 555 conjugate is very useful for immunohistochemistry experiments when the more commonly used mouse or rabbit primary/secondary antibodies are not feasible.

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