In single-cell sequencing, can we first perform primary cell culture on the tissue and then send it for testing after the cells have grown to a certain quantity?
In single-cell sequencing experiments, directly isolating cells from the original tissue is usually the best choice, rather than submitting them after primary cell culture. This is mainly because cell culture may introduce the following issues, affecting the accuracy and reliability of single-cell sequencing results:
- Culture may lead to loss of cellular heterogeneity.
- Culture conditions can alter the gene expression profile of cells.
- Increase the risk of contamination and background noise.
Recommended approach:
1. Preferably isolate single cells directly from tissue
(1) Obtain a single-cell suspension through enzymatic digestion or mechanical separation.
(2) Perform sequencing immediately to prevent changes in cell state.
2. Short-term culture under special circumstances
(1) If culture is necessary, limit the duration to 24-48 hours and carefully optimize culture conditions.
(2) Collect samples before and after culture to assess differences in gene expression.
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