The quantitative hard Surface time-kill Test method is excellent for use by disinfectant product developers because it is fast, relatively inexpensive, and provides information regarding the contact time that will be necessary to pass AOAC hard surface efficacy tests, such as the AOAC Germicidal Spray Products Test. Below, Microchem Laboratory presents a summary of the quantitative hard surface time-kill test, which is based on ASTM E1153, along with some of its strengths and weaknesses.
Summary of the Hard Surface Time-Kill Test

A microbial culture is prepared. For most bacteria, a 24 hour culture in nutrient broth works well. For most fungi, a spore preparation from a saline wash works well.

A volume of microbial culture (usually 0.010 ml to 0.020 ml) is placed onto the center of each of a number of sterile test surfaces. This inoculum can be spread over the sterile test surface in a circular pattern to achieve a thin, uniform coverage with the test microorganism if desired.

To measure initial microbial concentrations, one or more untreated, inoculated test surfaces are harvested and microorganisms are enumerated.

The remaining inoculated test surfaces are treated with the test product, each for a different length of time.

Immediately after the treatment times have elapsed, the test surfaces are placed into a solution that neutralizes the disinfecting action of the product, and microorganisms surviving treatment with the disinfectant or sanitizer are cultured and enumerated.

Results of the time-kill experiment are tabulated and reported, usually by charting microbial concentrations on the test surfaces as a function of treatment time with the disinfectant or sanitizer.

Strengths of the Hard Surface Time-Kill Test

The dried film of microorganisms used for this test makes the experimental model realistic, since typical "wet" vehicles of microbial surface contamination (e.g. mucous, blood, etc) will naturally dry onto surfaces over time.

Surface-based time-kill tests are relatively inexpensive.

The test parameters for surface-based time-kill tests are fairly easy to control in the laboratory setting, so valid comparisons can be made between various disinfectant and sanitizer products tested under the same conditions.

Weaknesses of the Hard Surface Time-Kill Test

The character of the test surface can have important bearing on the test outcome (e.g., porous surfaces are more difficult to disinfect than non-porous surfaces).

Environmental parameters related to the drying of the microbial inoculum can impact the test (e.g., low relative humidity will cause the microbial inoculum to dry more quickly than an inoculum dried at high relative humidity).

Some microorganisms tolerate drying better than others, which can make standardization of the dry microbial inoculum across microbial species challenging.

Microchem Laboratory is proud to be the premier research and development laboratory for disinfectants and sanitizers. The surface time-kill test is just one of many tools we use to help companies develop disinfectants and other antimicrobial products.

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