Sox2 Localization in Larval Xenopus Corneas

November 21, 2013

Cell and Developmental Biology
University of Illinois at Urbana-Champaign
Senior Research Specialist

Overall

Quality of Results

Ease-of-Optimization

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Company:

Aviva Systems Biology

Product Name:

Sox2 antibody

Catalog Number:

ARP31737

The goal of this research was to use known markers of pluripotency and examine their localization in the larval Xenopus cornea during lens regeneration. This antibody was chosen because of its similarity to the Xenopus Sox2 gene sequence.

Experimental Design and Results Summary

Applications

Immunohistochemistry

Sample

Xenopus laevis stage 50-52 cornea epithelium from control and regenerating specimens

Primary Incubation

Anti-Sox2 (Aviva Systems Biology ARP31737) overnight at 4 degrees Celcius (1:300 in blocking agent)

Blocking Agent

0.5% Triton/1X PBS/10% goat serum

Secondary Incubation

Goat-anti-mouse Alexafluor 546 (Molecular Probes); incubate overnight (1:300) at 4 degrees Celcius or 2 hours at RT

Tertiary Incubation

None

Detection

Fluorescence imaging

Results Summary

Control larval corneas yielded weak, but widespread cytoplasmic staining in both layers of the cornea epithelium. Additionally a small subset of cells located mainly in the basal epithelium displayed intense labeling around the periphery of the cornea. A dramatic increase of Sox2 labeling was observed in corneas where the lens had been removed 4 hours prior. These Sox2 labeled cells were present across the entire cornea, including the region overlying the pupillary space.

Additional Notes

Specimens gave the best results when fixed in 4% formaldehyde diluted in 1X PBS. Fixation times that worked varied from 1 hour to overnight at 4 degrees Celcius.

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Summary

The Good

Antibody was easily optimized for our system.

The Bad

Some background labeling was observed and was not suitable for Western blot analysis.

The Bottom Line

This antibody detected Sox2 labeled cells in Xenopus cornea samples. The background labeling issue could be averted with additional washes after the primary and secondary labeling steps.

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