ATP Assay Kit

March 12, 2015

University of South Florida
CMMB
Graduate Assistant

Overall

Quality of Results

Ease-of-Optimization

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Company:

BioVision

Product Name:

BioVision ATP Assay Kit

Catalog Number:

K354-100

The aim of the research was to quantify ATP levels in a mutant strain compared to the wild-type strain. We hypothesized that the mutant strain produced less ATP than the wild-type strain. This kit was selected for the ease of use, and the ability to quantify ATP levels with a plate reader that is routinely used in the lab.

Experimental Design and Results Summary

Application

Quantify ATP Levels

Starting Material

Bacterial cells

Protocol Overview

1. Cells were grown and lysed using a standard lysis method for cells (lysostaphin treatment for an hour). 2. Cells were centrifuged, and supernatant was stored on ice for the rest of the protocol. 3. Reaction mixes were set up according to the manufacturer's protocol using 50 ul of cell supernatant. 4. Reactions were incubated for an hour, not 30 minutes, in the dark. 5. OD 570 nm was measured with a plate reader. 6. Calculations for ATP were determined using a standard curve, that was set up alongside the experimental reaction mixes.

Tips

It is best to use concentrated cell samples if running the colorimetric assay, since it is less sensitive than the fluorometric readings.

Results Summary

We were able to determine lower levels of ATP in our mutant cells compared to our wild-type cells. This was expected as we predicted that our mutants were deficient in producing components of the electron transport chain.

Additional Notes

It would be helpful to have more than 100 assays as you easily run through them when optimizing volumes of cell lysates to use.

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Summary

The Good

The kit is easy to use, and manufacturer's protocol is easy to follow.

The Bad

The reagents do not store well for very long once opened.

The Bottom Line

I would recommend the use of the product. It would be improved with mores assays (to better optimize the protocol for specific needs). It would also be helpful if the reagents kept for longer so that the kit could be used for a longer period of time.

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