GB/T 11446.10-1997 Filter culture test method for total bacterial count in electronic grade water
Some standard content:
GB/T11446. 311446. 10—1997
GB/T11446-311446.10—1997 is a revision of GB11446.3—89 "General Rules for Testing Methods of Electronic Grade Water", GB11446.4—89 "Test Method for Resistivity of Electronic Grade Water", GB11446.5—89 "Atomic Absorption Spectrophotometric Test Method for Trace Metals in Electronic Grade Water", GB11446.6—89 "Spectrophotometric Test Method for Amount of Silica in Electronic Grade Water", GB11446.7—89 "Ion Chromatographic Test Method for Trace Nitrogen Ions in Electronic Grade Water", GB11446.8--89 "Test Method for Total Organic Carbon in Electronic Grade Water", GB11446.9—89 "Instrumental Test Method for Particles in Electronic Grade Water", and GB11446.10—89 "Filter Culture Test Method for Total Bacteria in Electronic Grade Water".
I Added technical indicators of metal nickel, nitrate ion, phosphate and sulfate ion to GB/T11446.1, and added "Atomic absorption spectrophotometry for determination of metal nickel and ion chromatography for determination of nitrate, phosphate and sulfate" to this standard. The test method for bacteria only uses the filter membrane culture method, and the method for measuring total carbon has been completely rewritten. The general rules for test methods, the method for measuring resistivity, and the method for determining total silicon have been revised and rewritten. From the date of implementation, this standard will replace GB11446.3~114 46.10—89. This standard was proposed by the Ministry of Electronics Industry of the People's Republic of China. This standard is under the jurisdiction of the Standardization Institute of the Ministry of Electronics Industry. The drafting units of this standard are: Institute of Semiconductors, Chinese Academy of Sciences. Institute of Standardization of the Ministry of Electronics Industry. The main drafters of this standard are: Wen Ruimei, Li Xiaoying, Wang Zaizhong, Xu Xuemin, Sun Ripan, Liu Renzhong, Xu Xiuxin. ..com1 Scope
National Standard of the People's Republic of China
Test method for total bacterial count in electronic grade water by membrane culture GB/T11446.10—1997
Test method for total bacterial count in electronic grade water by membrane filters This standard specifies the test method for the total bacterial count (live bacteria) in electronic grade water by membrane filters This standard is applicable to the measurement of the total bacterial count in electronic grade water of grade I to II. 2 Referenced standards
Replacement of GB11446.10--89
The following standards contain provisions that constitute the provisions of this standard by reference in this standard. When this standard is published, the versions shown are valid. All standards will be revised, and parties using this standard should explore the possibility of using the latest versions of the following standards. GB/T11446.1—1997 Electronic grade water
GB/T11446.31997 General rules for testing methods for electronic grade water 3 Definitions
3.1 Bacteria operation Bacteria operation Operations performed under conditions where no bacteria exist. When determining bacteria in water, the instruments, equipment and experimental equipment used must be sterilized in advance to obtain accurate results. 3.2 Bacterium colonies colony
The bacteria in water are so tiny that they cannot be observed with the naked eye. After a single bacterial body or spore is cultured on a solid medium for a certain period of time, it will grow and multiply to form a microbial group visible to the naked eye, which is a colony. 3.3 Bacterium culture method The quantitative water sample is filtered through an ultrafiltration membrane, and then the bacteria remaining on the surface of the ultrafiltration membrane are cultured with an appropriate culture medium to form colonies, which are then counted to measure the number of live bacteria in the water.
4 Principle
The bacteria in the water are filtered through a 0.45 μm microporous membrane and retained on the filter membrane. The dense membrane is attached to the substrate (filter paper that has absorbed TGY culture medium) and cultured under suitable conditions. The bacteria multiply into colonies visible to the naked eye. Dehydrogenase in bacteria reacts with 2,3,5-triphenyltetrazolium chloride (abbreviated as T.I.C) in the cell to make the bacteria red, increasing the contrast, which is used to measure and count. 5 Reagents
5.1 Blank water, 1st grade electronic grade water, sterile for 15 minutes at 1.4×10°Pa pressure in an autoclave. 5.2 IGY medium: 10.0g trypsin Chen (B3.R), 6.0g beef (BR), 2.0g glucose (AR) in 1L blank water, heat to dissolve. Cool to room temperature, adjust pH to 7.4~7.6 with 1 mol/L sodium hydroxide. Add 15.0g agar, heat to dissolve and filter while hot (with four layers of gauze and one layer of absorbent cotton as the medium), put in a 300 ml conical flask, sterilize and set aside. Approved by the State Administration of Technical Supervision on September 1, 1997, and implemented on September 1, 1998
GB/T 11446.101997
5.31 mol/L sodium hydroxide solution: 20 sodium hydroxide (A, R) dissolved in 500 ml of blank water, stored in a polyethylene bottle. 5.4 0.1% (W/V) TTC solution sterilized for use. 6 Instruments and equipment
6.1 Sterile room (or sterile workbench): equipped with a UV lamp with a power of 2 W/m or more, placed 1.5 m from the workbench. 6.2 Electric constant temperature incubator: (20~60)11℃. 6.3 High pressure steam sterilizer.
6.4 Constant temperature oven,
6.5 Stereo microscope.
6.6 Microporous organic filter membrane: ±50mm, pore size 0.45μm grid membrane. 6.7 Filter $50mm cup-type stainless steel filter. 6.8 Various glassware blood bottles, angle bottles, culture blood, measuring cylinders, etc. 6.9 Alcohol lamp, flat-head tweezers, gauze, cotton, qualitative filter paper, etc. 6.10 Filtration system: as shown in the figure below
1—cup, 2—filter membrane, 3—membrane mesh 14-single filter bottle 5 buffer bottle ±6 Vacuum pump filtration system diagram
7 Analysis steps
7.1 Preparation before traceability
7.1.1 Open the sterile room and purification workbench for 1h~2h to make it enter a stable working state. 7.1.2 Sterilization of utensils: Wash the culture blood, sampling bottles and other glassware with electronic grade water, dry them, wrap them with paper, place them in a lead special container and put them in a constant temperature box (160℃～165℃>: heat sterilize for 2 hours. 7.1.3 Sterilization of organic filter membranes: Boil them in electronic grade water for 15 minutes, add new water after boiling water, and repeat the process three times. Sterilize them under ultraviolet light for 1 hour to 2 hours. 7.2 Sedimentation of water samples
Take 2.0 mL of TTC solution in 100 ml of culture medium, mix it in the culture medium (about 3 mm~4 mm thick), and cool it for use. Take an appropriate amount of water sample after operation: 2 1L of water, filter it through the filter device, rinse it with 5L of blank water, and draw out. Use tweezers to place the filter membrane on the prepared culture medium, with the membrane facing up, cover it with a lid, and wait for it to solidify. Then turn it over and put it in the incubator. Keep it in a constant temperature and dark at 37℃ for 48h, then take it out for observation and counting of colonies under a microscope. Follow the same steps to carry out a blank experiment. Each sample is measured three times in parallel, and the average value is taken. 8 Calculation of analysis results
The total number of bacteria in water is calculated by the following formula:
GB/T11446.10—1997
Where: A——Total number of bacteria per milliliter of water sample, pieces,
——Number of colonies on the filter membrane, pieces;
——Number of colonies in the blank experiment, pieces
V—Volume of the tested water sample, mL.
Test report
According to GB/T 11446.3—1997 Chapter 6. Precautions bzxz.net
10.1 Special attention should be paid to the contamination of foreign microorganisms in bacterial analysis. 10.2 When placing the bacterial film on the culture medium, there should be no gaps or bubbles in the middle. 10.3 During the heating process of preparing the culture medium, the lost water should be replenished.
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