Worked Well On Paraffin Embedded Mouse Brain Sections

July 20, 2017

Department of Cell Biology
SUNY Downstate Medical Center
PhD Candidate (Graduate Student)

Overall

Quality of Results

Ease-of-Optimization

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Company:

Abcam

Product Name:

A goat anti-rabbit IgG H&L DyLight® 550

Catalog Number:

ab96884

We study how cobalamin deficiency alters learning and memory using a KO mouse model. The KO mouse showed impairment in the early expression of long-term potentiation (LTP). In order to determine the underlying mechanism of this impairment, we studied the expression of postsynaptic domain proteins as well as AMPA receptors in the hippocampus of the KO mouse. We determined the expression by IHC-P, using DyLight 550 as the secondary Ab.

Experimental Design and Results Summary

Applications

Immunohistochemistry

Sample

Paraffin embedded mouse brain sections

Primary Incubation

1:200, 22 hrs at 4 degree C

Blocking Agent

Normal goat serum

Secondary Incubation

1:200, 16 hrs at 4 degree C

Tertiary Incubation

NA

Detection

Fluorescence

Results Summary

We got fluorescence (represneting the expression of the proteins) as particles that we could count to quantify the amount of protein. This was used for comparing the expression levels between control and the KO mouse.

DOI or PMID #

28545069

Additional Notes

Antigen retrieval was performed prior to immunostaining

Image Gallery

Summary

The Good

Highly specific with no background even with lower dilution

The Bad

Gives fluorescence with multiple filters

The Bottom Line

Works wonderfully on mouse sections. We tried immunostaining with another secondary Ab that was HRP conjugated. It gave simply dark color looking like a background whereas, DyLight 550 gave microdots (particles) as the expression of a protein that we could quantify. It gave fluorescence with multiple filters, however, we used a simple Olympus fluorescence microscope. This issue might not occur in confocal imaging.

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