Anaerobic microbial hungate rolling tube method

Hungate Hungate anaerobic tube culture technology can be used not only for the separation of beneficial anaerobic bacteria such as bifidobacteria, but also for the counting of live bacteria, as well as harmful spoilage bacteria (such as butyric acid bacteria) or pathogenic bacteria (such as botulinum Isolation and identification of Bacillus licheniformis.
The Hungate Hungate Anaerobic Rolling Tube Technology was an anaerobic culture technique first proposed by American microbiologist Hungate in 1950 and applied to rumen anaerobic microbiology . In the future, this technology has undergone continuous improvement over the past few decades, which has made Hengate's anaerobic technology more and more perfect, and has gradually developed into a complete set of technologies for studying anaerobic microorganisms . And years of practice have proven it to be an extremely effective technique for studying rigorous, obligate anaerobic bacteria.
1. Copper column system The oxygen removal copper column is a hard glass tube with copper wire or copper chips inside. The tube is 40-400 mm in size, the two sections are processed into a funnel, the outer wall is surrounded by a heating belt, and connected to a transformer to control the voltage and stabilize the temperature of the copper column. The two ends of the copper column are connected with a hose, one end is connected to the gas cylinder, and one end is connected to the outlet port. Since gases from gas cylinders such as N2, CO2 and H2 usually contain O2, when these gases pass through a copper column with a temperature of about 360 °C, a small amount of O2 in copper and gas combines to form CuO, and the copper column is bright. Yellow turns black. When H2 is introduced into the oxidized copper column, the oxygen in H2 and CuO combine to form H2O, and CuO is reduced to copper, and the copper column is bright yellow. This copper column can be used repeatedly and continuously for oxygen removal purposes. Of course, the H2 source can also be generated by a hydrogen generator.
5.2 Pre-reduction medium and dilution preparation When pre-reducing medium and diluent are prepared, the prepared medium and diluent are first boiled and oxygenated, and then semi-quantitatively loaded with a semi-quantitative sampler to the screw anaerobic In the test tube , the general agar medium was loaded with 4.5-5 ml, the diluted solution was loaded with 9 ml, and a long needle with N2 gas was inserted to exclude O2. At this point, it can be clearly seen that the redox indicator added to the medium - resazurin from blue to red, finally becomes colorless, indicating that the test tube has become anaerobic, and then the butadiene rubber plug of the screw is covered and Screw cap, sterilized for use.
5.3 Preparation of different dilutions of bifidobacteria samples Accurately weigh 1 g of solid under sterile conditions or take 1 ml of a homogeneous liquid sample with a sterile syringe, then add to an anaerobic test tube containing pre-reduced physiological saline, using an oscillator Shake it evenly to make a 10-1 dilution. A 1 ml dilution of 10-1 was pipetted into a separate tube containing 9 ml of physiological saline using a sterile syringe to make a 10-2 dilution. According to this method, serial dilutions of 10 times to 10-7 were sequentially performed to prepare sample dilutions of different concentrations. Roller culture counts are usually performed at three dilutions of 10-5, 10-6, and 10-7.
5.4 Anaerobic Rolling Tube Culture Method <br> Place the test tube containing the sterile sterile oxygen-free agar medium in a constant temperature water bath at about 50 °C, and draw 10-5, 10-6, 10-1 with a 1 ml sterile syringe. 7 ml of each of the three dilutions of 0.1 ml in a melted agar medium screw anaerobic test tube , and then flatly placed in a tray containing ice cubes or a special rolling machine to quickly roll, so that the bacteria The thawing medium is immediately solidified into a thin layer on the inner wall of the screw anaerobic tube . Each dilution was repeated 3 times and then cultured in a 37 ° C (yoghurt sample 42 ° C) incubator. After 24 to 48 hours of incubation, the colonies visible to the naked eye can be grown in the agar layer or on the surface of the screw anaerobic test tube .

Anaerobic tubes are an important part of the Hengate anaerobic tube technology and are designed for the cultivation of anaerobic microorganisms.
Screw anaerobic test tube http://?product-98.html
Anaerobic culture tube (aluminum sealed type) http://?product-97.html