常见培养基质量控制问题汇总
发布时间：
2022-12-24
作者：
上海微生物培养基
常见培养基质量控制问题汇总
1、平板涂布和平板划线培养基表面水分过多，菌落蔓延如何解决？
对于采用表面接种形式培养的固体培养基，应先对琼脂表面进行干燥：揭开平皿盖，将平板倒扣于烘箱或培养箱中（温度设为25℃～50℃）；或放在有对流的无菌净化台中，直到培养基表面的水滴消失为止。注意不要过度干燥。商品化的平板琼脂培养基应按照厂商提供的说明使用。
2、灭菌后的培养基的存放时间，保存方式？
琼脂培养基的融化
将培养基放到沸水浴中或采用有相同效果的方法（如高压锅中的层流蒸汽）使之融化。经过高压的培养基应尽量减少重新加热时间，融化后避免过度加热。融化后应短暂置于室温中（如2 min）以避免玻璃瓶破碎。
融化后的培养基放入47℃～50℃的恒温水浴锅中冷却保温(可根据实际培养基凝固温度适当提高水浴锅温度)，直至使用，培养基达到47℃～50℃的时间与培养基的品种、体积、数量有关。融化后的培养基应尽快使用，放置时间一般不应超过4 h。未用完的培养基不能重新凝固留待下次使用。敏感的培养基尤应注意，融化后保温时间应尽量缩短，如有特定要求可参考指定的标准。
倾注到样品中的培养基温度应控制在约45℃左右。
平板的制备和储存
倾注融化的培养基到平皿中，使之在平皿中形成厚度至少为3 mm（直径90 mm的平皿，通常要加入18 mL～20 mL琼脂培养基）。将平皿盖好皿盖后放到水平平面使琼脂冷却凝固。如果平板需储存，或者培养时间超过48 h或培养温度高于40 ℃，则需要倾注更多的培养基。凝固后的培养基应立即使用或存放于暗处和（或）5℃±3℃冰箱的密封袋中，以防止培养基成分的改变。在平板底部或侧边做好标记，标记的内容包括名称、制备日期和（或）有效期。也可使用适宜的培养基编码系统进行标记。
将倒好的平板放在密封的袋子中冷藏保存可延长储存期限。为了避免冷凝水的产生，平板应冷却后再装入袋中。储存前不要对培养基表面进行干燥处理。
对于采用表面接种形式培养的固体培养基，应先对琼脂表面进行干燥：揭开平皿盖，将平板倒扣于烘箱或培养箱中（温度设为25℃～50℃）；或放在有对流的无菌净化台中，直到培养基表面的水滴消失为止。注意不要过度干燥。商品化的平板琼脂培养基应按照厂商提供的说明使用。
Summary of common media quality control problems
1. How to solve the problem of colony spread when there is too much water on the surface of plate coating and plate marking media?
For solid medium using surface vaccination form, should be carried out on the AGAR surface drying: uncover plate cover, will buckle the tablet in the oven or in the incubator (temperature is set to 25 ℃ to 50 ℃); Or put it in a sterile purification stand with convection until the droplets on the surface of the culture medium disappear. Be careful not to overdry. The commercialized plate AGAR media shall be used in accordance with the instructions provided by the manufacturer.
2. Storage time and preservation method of sterilized media?
Melting of AGAR medium
Melt the medium in a boiling water bath or by using the same method as laminar steam in a pressure cooker. The high pressure medium should minimize the reheating time and avoid excessive heating after melting. After melting, the glass bottle should be placed at room temperature (eg 2 min) for a short time to avoid breakage.
Melted medium in 47 ℃ to 50 ℃ constant temperature water-bath water in cooling heat preservation (according to actual medium solidification temperature raise the temperature of water bath pot), until the use, medium to 47 ℃ ~ 50 ℃ time associated with the variety, size, quantity of medium. The melted medium should be used as soon as possible and generally should not be placed for more than 4 hours. The unused medium cannot be re-solidified for another time. Especially for sensitive media, it should be noted that the time of heat preservation after melting should be shortened as far as possible. If there are specific requirements, the specified standards can be referred to.
Poured into the sample medium temperature should be controlled at about 45 ℃ or so.
Plate preparation and storage
Pour the melted medium into the plate to form a plate at least 3 mm thick (90 mm in diameter, usually 18 mL ~ 20 mL AGAR medium). Cover the plate and put it on the horizontal plane to cool the AGAR. If tablet should be stored, or more than 48 h of incubation time or the temperature higher than 40 ℃, then you need to devote more of the culture medium. After solidification of media should be used immediately or stored in the dark and (or) 5 ℃ + 3 ℃ refrigerator sealing bag, in order to prevent the medium components change. Mark the bottom or side of the plate with a name, preparation date, and/or expiration date. The appropriate medium coding system can also be used for labeling.
Keep the inverted plate in a sealed bag and refrigerate for longer storage. In order to avoid condensation, the plate should be cooled and then put into a bag. Do not dry the surface of the medium before storage.
For solid medium using surface vaccination form, should be carried out on the AGAR surface drying: uncover plate cover, will buckle the tablet in the oven or in the incubator (temperature is set to 25 ℃ to 50 ℃); Or put it in a sterile purification stand with convection until the droplets on the surface of the culture medium disappear. Be careful not to overdry. The commercialized plate AGAR media shall be used in accordance with the instructions provided by the manufacturer.