GABA Antibody (Rabbit Anti-Mouse) for Immunostaining Mouse Brain Slices

January 18, 2016

Biomedical Engineering
Boston University
Graduate Student

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Company:

Sigma-Aldrich

Product Name:

Anti-GABA antibody produced in rabbit

Catalog Number:

A2052

Browse Similar Products: GABA Immunohistochemistry Antibodies

I have used this antibody hundreds of times in the past two years to immunostain mouse and rat brain sections and to identify inhibitory neurons (and colocalization with an EGFP reporter). This antibody always stains well with low background in both the cortex and hippocampus, the two brain regions I am concerned with.

Experimental Design and Results Summary

Applications

Immunohistochemistry

Sample

Fixed mouse and rat brain slices (40 um)

Primary Incubation

1:1000 in PBS with 0.5% Triton X-100 and 2% goat serum (500 uL per brain slice). Incubated overnight (~16 hours) gently rocking at 4 degrees C.

Blocking Agent

2% goat serum in PBS with 0.5% Triton X-100

Secondary Incubation

Alexa Fluor 568 goat anti-rabbit, 1:1000 in PBS with 0.5% Triton X-100 and 2% goat serum (500 uL per brain slice). Incubated 2-3 hours at room temperature, gently rocking.

Tertiary Incubation

ToPro-3, 1:5000 in PBS with 0.5% Triton X-100 (500 uL per brain slice). Incubated 30 minutes at room temperature, gently rocking

Detection

Confocal microscopy with Olympus FluoView FV1000 software

Results Summary

GABA neurons were easily identifiable with this antibody, which enabled manual quantification of colocalization between my EGFP construct (built to target inhibitory neurons) and the inhibitory neuron population in the rodent neocortex and hippocampus.

Additional Notes

I repeatedly froze and thawed this antibody multiple times and it continued to give great staining results.

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Summary

The Good

Excellent GABA neuron labeling and low background in fixed brain tissue

The Bad

There was oversaturation of signal at the top and bottom ~5 um of each brain slice

The Bottom Line

Great antibody for imaging inhibitory (GABA) neurons in mouse and rat brain slices, especially in neocortex and hippocampus

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