细胞培养技术与其他一般实验室工作的主要区别在于要求保持无菌操作，Avoid the influence of microorganisms and other harmful factors。A general standard cell culture chamber should include a preparation chamber、Preparation room and culture room。The three chambers are both interconnected and relatively independent，Each completes different operations in the culture process。
【实验目的】
①了解培养室的设置和设备。
(2) Learn the concept of aseptic and the essentials of aseptic operation。
【操作步骤】
1．实验室设置
（1）配液室
（2）准备室
（3）培养室
培养室内要完全密闭，Maintain a constant temperature，The following points should be paid attention to in the design：
(1) The location of the culture room is best located on the shaded side，Places where direct sunlight cannot be reached，Prevent the indoor temperature from rising。
(2) The height of the ceiling should not exceed 2 meters，In order to ensure the effective sterilization effect of the UV lamp，
(3) The door is generally sliding door，to prevent air movement。
(4) Ceiling、The floor and surrounding walls should be smooth and free of dead corners，To tile or paint，It's designed like this，First, it is easy to clean and disinfect，In addition, it is not easy to accumulate dust in the corners of the wall。
2．实验室常用设备
（1）准备室的设备
①双蒸馏水蒸馏器：Prepare double distilled water。
(2) Acid tank：Hold lotion，Soak glassware。
(3) Oven：Washed glassware is oven dried。
(4) Pressure cooker：glassware、Dissecting utensils、Partial liquid、Sterilization of plastic utensils, etc。The effective sterilization pressure and time are different for different items。
(5) Locker 1：Place unsterilized items。
(6) Locker 2：Place sanitized items。
(7) Packing table：For packaging before sterilization。
Preparation room precautions：
(1) Prevent the still from being roasted dry。
(2) Water seeping out from the placed pool。
(3) Do not splash acid on clothing or the ground。
(4) Do not block the pressure cooker exhaust valve and safety valve。
(5) Sterilized items should be stored separately from unsterilized items。
（2）配液室的设备
①天平（扭力天平和电子天平）：称量用
②pH计：
(3) Magnetic stirrer：
（3）细胞培养室的设备
①超净工作台：
There are many types of clean benches，There is a one-sided single、Single-sided double、Double-sided double or double-sided quadruple, etc，It works by utilizing a blower，Enables the air to pass through highly efficient air filtration，Slowly pass through the countertop，This results in a sterile and homogeneous air in the working environment。
Depending on the laminar flow method，Ultra-clean workbenches are mainly divided into two types: horizontal laminar flow type and vertical laminar flow type，The rationale is much the same，The indoor air is initially filtered through a coarse filter，It is pressed into the static pressure box by a centrifugal fan，Then pass through the high-efficiency air filter，The resulting clean air flow passes through the sterility at a certain uniform cross-sectional velocity，So as to form a dust-free and sterile high-cleanliness working environment。However, the direction of the airflow is different between the two types of stations。
There are several issues that should be paid attention to when using a clean bench：
A. The purification workbench is best installed in a dust-free room，It is best to be in a well-isolated sterile room，In order to avoid excessive dust, the filter will be easily blocked，Reduces the purification effect，Shorten the service life。
B. Newly installed or long-term unused workstations，The workbench and surroundings must be cleaned with a vacuum cleaner or a tool that does not produce fibers before work，Then use drug sterilization or ultraviolet sterilization for sterilization。
C. Every time the workbench is used，The countertop should be scrubbed with 75% alcohol first，And treat the microorganisms accumulated in the purification work area with 30~50min ultraviolet sterilization lamp in advance。After turning off the UV lamp, the blower should be started and let it run for two minutes before the incubation operation。
D. Unnecessary items should not be stored in the purification work area，to keep the clean airflow pattern undisturbed。
E. Generally，The HEPA filter is replaced every three years。Replacing the high-efficiency filter should be operated by a professional，to keep the seal well。The coarse filter should be removed and cleaned regularly，The time should be determined according to the cleanliness of the environment，It is usually done every 3~6 months。
F. Every time you use the purification table, you should remove the items on the work surface in time，And scrub the countertop with sprinkler to keep it clean at all times。
(2) 4 °C freezer and low temperature freezer：
③CO2培养箱：
A．一定浓度的CO2对细胞生长，In particular, primary culture and single-cell culture have a facilitating effect (5%)。
B．一定浓度的CO2可维持培养液恒定的pH。Suitable for open cultures，After the incubator is sealed，It can be cultivated in a general incubator，However, when using Petri dishes or multi-well plates，则需要高湿度和高CO2含量的空气。
C. It increases humidity，The box is equipped with a water bath，Humidity in the incubator can be guaranteed。
D. The incubator is equipped with a UV lamp。
E．通过气体流量计可以调节CO2和空气的比例。
F. Preservatives (dioxacetic acid) can be added to the water。可防止因CO2培养箱中湿度较高，Prone to mold。
(4) Ozone generator：
(4) Centrifuge：
⑤CO2钢瓶：
(6) Liquid nitrogen tank：
(7) Lockers：用来存放杂物
⑧紫外灯：
⑨空气净化系统
⑩倒置显微镜
3．无菌操作
（1）无菌室的灭菌：
(1) The ultraviolet lamp should be turned on when no one is there；
(2) Wear a sterile gown when entering the sterile room、Wear a hat and mask；
(3) Clean and disinfect once a week；
④所用物品要以外科手术方式严格消毒；
⑤室内台面要要保持洁净，做完实验后，用75％酒精或千分之三新洁尔灭棉球擦拭超净工作台的台内、边台的台面、倒置显微镜的载物台等；
⑥所用物品要一次性准备好；
⑦瓶口要用酒精火焰消毒；
⑧尽量减少出入。
（2）实验人员的无菌准备：
①肥皂洗手
②穿好隔离衣
③酒精棉球擦手
细胞培养室必备的设备有：
1. 超净台；2. 二氧化碳培养箱；3. 倒置显微镜；4. 二氧化碳压缩气体；5. 液氮罐；6.低温冰箱（-20度和-70度）；7. 过滤装置；8. 离心机；9. 酶标仪等等。
试剂和小的工器具也很多，as：培养基、培养瓶、培养板、移液枪、酸缸、滴管、离心管、血球计数板、DMO、PBS、胰酶等等。
细胞培养室，最好选择独立房间，超净台应放置在比较洁净的房间，如果条件允许，最好按洁净厂房要求设计细胞培养室，即使房间做不到空气过滤，也至少应是专用的，进出时要更换工作衣。