Microbial Viability Assay Kit-WST

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Microbial Viability Assay Kit-WST - M439

Microbial Viability Assay Kit-WST (500 tests) - M439-10

Item #	Description/Size	Availability	Qty Break	Price	Quantity
M439-10	500 tests	1-2 business days	1	$200.00

*Estimated. Exact shipping date will be notified.
For Research Use Only Products

Colorimetric microplate assay

Wide variety of microorganism detection

No harvesting or washing required

Description
Application
Data
Manual

Kit Contents

WST solution	1 ml x 5 tubes
Electron mediator reagent/ DMSO	0.5 ml x 1 tube

Storage Condition : 0-5ºC
Shipping Condition : ambient temperature

Required Equipment and Materials
plate reader with 450 nm filter; 96-well culture plate, 10 μl, 100-200 μl, and multi-channel pipettes; incubator

Product Description

Viable bacterial cell detections are very important for analyzing bacteria contamination in food or evaluating the cleanliness of facilities in order to protect us from food poisoning and infections. Bacterial cell detections are also used for the screening of sanitizing agents and drug resistance detections. Generally, counting the number of colonies on an agar plate is the standard method for determining the number of viable bacterial cells in samples. However, colony formations require one to several days. Dojindo’s Microbial Viability Assay Kit-WST can be used to determine the number of viable bacterial cells in a sample by a colorimetric method and can be applied to 96-well microplate assays. The electron mediator in the kit receives electrons from viable bacterial cells and transfers the electrones to WST, one of the water-soluble tetrazolium salts developed by Dojindo. Bacterial cell viability then can be determined by monitoring the color intensity of WST formazan dye. Since several types of media used for bacterial cell cultures and components do not interfere with the assay(Fig. 5), simply add the assay solution and incubate for one to several hours to determine the initial number of viable bacterial cells in the sample(Fig. 2). This assay kit was co-developed with the Biotechnology and Food Research Institute, Fukuoka Industrial Technology Center.

Fig. 1 Bacterial cell viability detection mechanism.

Assay Data

Fig. 2 Correlation between initial number of E. coli and time-dependent O.D. increase. The initial number of viable E.coli were determined by a colony counting method.

Fig. 3 Correlation between the initial number of SA and time-dependent O.D. increase. The initial number of viable SA were determined by the colony counting method.

Comparison Chart

Fig. 5 Influence of culture media or substances used for bacterial cell culture.
The data indicated that WST is less sensitive to various culture media or substances which are used for bacterial cell culture. WST is a better tetrazolium salt than XTT for bacterial cell viability assays.

General Procedure 2
Determination of the susceptibility of Staphylococcus aureus to oxacillin
Oxacillin: antimicrobial agent: 0-64 μg/ml
Microorganism: Staphylococcus aureus (SA)
Methicillin-resistant Staphylococcus aureus (MRSA)
1. Culture SA or MRSA with Mueller-Hintonmedium containing various concentrations of Oxacillin for 6 hours at 35ºC.
2. Add Microbial Viability Assay solution equal to 1/20 the volume of the culture medium.
3. Incubate for 2 hours at 35^ºC.
4. Measure the O.D. at 450 nm to determine the MIC (Minimum inhibitory concentration).

Fig. 4 Susceptibility test of SA and MRSA against Oxacillin.
The data indicated that MRSA has lower susceptibility than SA. The MICs of MRSA (32 μg/ml) and SA (0.5 μg/ml) are close to the MICs determined by the CLSI (Clinical and Laboratory Standards Institute) method.

Reference
1. T. Tsukatani, et al., Colorimetric cell proliferation assay for microorganisms in microtiter plate using water-soluble tetrazolium salts. J Microbiol Methods. 2008;75:109-116.
2. T. Tsukatani, et al., Colorimetric microbial viability assay based on reduction of water-soluble tetrazolium salts for antimicrobial susceptibility testing and screening of antimicrobial substances. Anal Biochem. 2009;393:117-125.

Appropriate Cell Density

Table 6 Initial cell number can reach O.D. 0.5 with 1-hour and 4-hour incubation.
The initial cell number of each microorganism was determined by colony counting. Each microorganism cell culture was diluted with medium and 190 μl of the cell culture was added to each well. Then 10 μl of assay solution was added. The cells were incubated at 30^oC or 37^oC for 1 hour and 4 hours to determine how many cells are required to reach O.D.=0.5 at 460 nm.