Sterility test method and totally enclosed bacterial ampoule incubator used by it

摘要

A sterility test method includes: selecting strain and culture medium, preparing bacterial cultures, transcribing fingerprint characteristics in thermograms as indices to verify the characteristics, drawing the thermodynamic parameters of the thermogram, determining the positive judgment index and performing sterility test for the samples. A fully-enclosed bacteria collecting ampoule incubator includes bacteria collecting ampoule system, sample and liquid feeding system and peristalsis liquid discharge system. The sample and liquid feeding system is connected with the bacteria collecting ampoule system by the liquid intake tube; and the bacteria collecting ampoule system is connected with the peristalsis liquid discharge system by the liquid drainage tube. The invention is characterized by short inspection time, high sensitivity, high automation and accurate test results on microbial contamination. It can also provide the overall process curve on the growth conditions. Such curve is provided with relatively favorable fingerprint, which enables qualitative analysis on the microbial contamination conditions.

主权项

1. A method for testing sterility of a sample comprising the steps of:
(1) preparing bacterial cultures comprising the steps of: cultivating different bacterial strains in a sterile culture medium to obtain bacterial cultures with different concentrations and different survival conditions as positive controls for recording fingerprint characteristic thermograms of the different bacterial strains; wherein the method for obtaining the bacterial cultures of different concentrations comprises: washing fresh bacteria cultures to obtain eluents, diluting the eluents with 0.9% sterile sodium chloride solution to produce a series of 10-fold dilutions; wherein the method for obtaining the bacterial cultures of different survival conditions comprises: filtering and eluting the bacterial cultures to obtain eluents, placing the eluents in a freezer at −70° C. and in a water bath at 60° C., respectively, for 2 h, and diluting the eluents with 0.9% sterile sodium chloride solution to produce a series of 10-fold dilutions; (2) recording the fingerprint characteristic thermograms of the different bacterial strains as diagnostic characteristics, wherein the recording comprises: placing the bacterial cultures obtained in Step (1) in a microcalorimeter under conditions suitable for growth of said bacterial cultures, recording the thermograms at different concentrations and under different survival conditions of the different bacterial strains to obtain the fingerprint characteristic thermograms of the different bacterial strains; (3) obtaining the thermodynamic parameters from the thermograms obtained in Step (2), and determining positive identification time indices for the bacterial strains; wherein the thermodynamic parameters comprise a time-dependent positive detection channel thermal power Pi and a corresponding blank control channel thermal power P0, the maximum thermal power Pmax, time required to reach the maximum thermal power Tmax, total heat production Htotal, and a slope k obtained from a 15-min segment on each exponential growth curve; and a positive identification time index for a strain obtained by recording the occurrence time for k≧0, which is an essential condition for determining microbial contamination of a sample, and establishing a positive identification time index for the growth of the microorganism; and (4) assessing sterility of the sample to be tested comprising the steps of: filtering the sample to be tested, rinsing products collected on the filter with a sterile solution; mixing the filtrate of the sample with the culture medium, putting the mixture into the detection channel of the microcalorimeter, recording a thermogram; comparing with the fingerprint characteristic thermograms for the different bacterial strains from Step (2) and the positive identification time indices for the different bacterial strains from Step (3), and determining the microbial contamination of the tested sample using the positive identification time indices for the growth of the microorganism in Step (3) as follows:
Td=Time[(Pi−P0)/|P0|≧3] wherein the time duration where the difference between the thermal power of the detection channel Pi and the thermal power P0 is greater than three times of the absolute value of P0 of the blank channel, taken as the detection time point (Td), indicates microbial contamination of the sample.