The quality of cell samples directly affects the reliability and reproducibility of experimental results. Shanghai Keborui Biotechnology will systematically introduce the core points of cell sample preparation and preservation, providing standardized operational references for researchers to ensure that cell samples are preserved in their optimal state, maintaining their biological characteristics and experimental value.
1、 Key steps in cell sample preparation
1. Preparation before Preparation
-Clear experimental design: Determine the preparation plan based on the subsequent experimental objectives (such as proteomics, transcriptomics, cell culture, etc.)
-Preparation of reagents and consumables: All reagents and consumables that come into contact with cells must be sterile and free of nucleases/proteases contamination
-Equipment pre cooling: Centrifuge rotors, freezing tubes, etc. are pre cooled to an appropriate temperature in advance
2. Cell collection and processing
-Cell status confirmation: Ensure that the cells are in logarithmic growth phase and the live cell rate is greater than 95%
-Gentle treatment: Avoid severe blows and wash with pre cooled PBS or saline solution
-Timely handling: Handle as soon as possible after removal from the incubator to reduce cellular stress response
3. Special handling requirements
-Protein analysis sample: Add protease inhibitor mixture to avoid repeated freeze-thaw cycles
-Nucleic acid analysis sample: Use RNase/DNase free consumables and add RNA protectant
-Metabolomics samples: Rapid quenching of metabolic activity using pre cooled methanol or specialized quenching solution
2、 Cell preservation methods and selection
1. Short term storage (<24 hours)
-4 ℃ storage: suspended in complete culture medium or buffer solution
-Applicable situations: Flow cytometry, live cell imaging and other experiments conducted on the same day
-Caution: Avoid bacterial contamination and store in a sealed container
2. Mid term storage (1 week to 1 month)
--80 ℃ storage: Add suitable cryoprotectant
-Recommended frozen storage solution:
-Cell culture: fetal bovine serum containing 10% DMSO
-Protein analysis: lysis buffer containing protease inhibitors
-Nucleic acid analysis: RNA protection reagents (such as TRIzol or specialized preservation solution)
3. Long term storage (>1 month)
-Liquid nitrogen preservation (-196 ℃): the most effective method for maintaining cell viability
-Freezing program:
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1. Program cooling: 4 ℃ for 30 minutes → -20 ℃ for 2 hours → -80 ℃ overnight
2. Transfer to liquid nitrogen gas phase (-150 ℃) for long-term storage
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-Freezing tube selection: label cell information, freezing date, passage times, operator
3、 Key points of quality control
1. Cell viability testing
-Perform trypan blue staining or MTT assay before and after cryopreservation
-After recovery, cell viability should be>80% (ideally>90%)
2. Pollution control
-Regularly detect mycoplasma contamination
-Visual observation and microscopic examination of bacterial and fungal contamination
-Cell cross contamination STR identification
3. Marking and Recording
-QR code or barcode management system
-Detailed records of cell sources, processing procedures, and storage locations
4、 Common problem solutions
|Problem | Possible Cause | Solution|
|Low cell activity after recovery | Rapid cooling during cryopreservation | Use of programmed cooling box|
|Protein degradation | Insufficient protease inhibitors | Adding a mixture of multiple protease inhibitors|
|RNA degradation | RNase contamination during operation | Using RNase free consumables for rapid processing|
|Mycoplasma contamination | Primary cell or operational contamination | Regular testing and use of mycoplasma scavengers|
5、 Safety precautions
1. Biosafety: Comply with the corresponding biosafety level requirements based on the cell source
2. Liquid nitrogen operation: Wear a protective face mask and gloves to prevent frostbite
3. Chemical safety: DMSO and other reagents should be handled properly to avoid direct contact with the skin
4. Waste disposal: Cell waste is processed according to the biological hazardous material disposal process
6、 Summary of recommended saving schemes
|Application direction | Recommended storage method | Storage temperature | Expected storage time|
|Cell Resuscitation Culture | DMSO+Serum Cryopreservation | Liquid Nitrogen | Over 10 Years|
|Western Blot | RIPA lysate | -80 ℃ | 1 year|
|RNA extraction | TRIzol or RNA protectant | -80 ℃ | 6 months|
|Metabolomics | 80% methanol quenching | -80 ℃ | 3 months|
|Flow cytometry | PBS containing 2% FBS | 4 ℃ | 24 hours|
7、 Intelligent management system
Suggest establishing an electronic management system for cell samples in the laboratory, including:
-Real time tracking of sample location
-Automatic update of freezing/retrieval records
-Regular inventory checks and reminders
-Quick retrieval of sample information

High quality cell samples are the cornerstone of successful experiments. By standardizing the preparation process and scientific preservation methods, the biological integrity of cell samples can be maximally maintained. With the development of new technologies such as single-cell analysis and spatial transcriptomics, the demand for cell sample quality will further increase. Establishing strict sample management standards is not only necessary to ensure the reliability of current experiments, but also a strategic investment to preserve valuable biological resources for future research.