1 / 11

BIOT 2922

BIOT 2922. Tissue Culture Methods. Objectives. Work aseptically. Maintain mammalian cells in healthy, uncontaminated state for 2-4 weeks. Manipulate mammalian cells. Cell lines. Chinese hamster ovary (CHO) cells Introduced in the 1960s. Used in a cultured monolayer in culture flasks.

tan
Download Presentation

BIOT 2922

An Image/Link below is provided (as is) to download presentation Download Policy: Content on the Website is provided to you AS IS for your information and personal use and may not be sold / licensed / shared on other websites without getting consent from its author. Content is provided to you AS IS for your information and personal use only. Download presentation by click this link. While downloading, if for some reason you are not able to download a presentation, the publisher may have deleted the file from their server. During download, if you can't get a presentation, the file might be deleted by the publisher.

E N D

Presentation Transcript

  1. BIOT 2922 Tissue Culture Methods

  2. Objectives • Work aseptically. • Maintain mammalian cells in healthy, uncontaminated state for 2-4 weeks. • Manipulate mammalian cells.

  3. Cell lines • Chinese hamster ovary (CHO) cells • Introduced in the 1960s. • Used in a cultured monolayer in culture flasks. • Used in studies of genetics, toxicity screening, nutrition and gene expression particularly expression of recombinant proteins. • Mouse myeloma cells • Cancer cell line. • Grow in clumps.

  4. CHO cells

  5. Myeloma cells

  6. Working with cell lines • Each student has own media and cells. • Make complete and incomplete media and test for sterility. • Obtain cells/monitor cell growth. • Use of inverted microscope. • Prepare laminar flow hood. Monitor CO2 incubator (Fyrite procedure for level). • Handle cells, split cells, count cells, freeze down cells.

  7. Laminar Flow Hood

  8. CHO cells 10% confluent ~100% confluent Confluency: coverage or proliferation that the cells are allowed over or throughout the culture medium

  9. First Day • 2 students make 1 L incomplete media, split into two 500 mL bottles. • Prep hood. • Each student makes 50 mL complete media (do this in hood). • Test both for sterility (Fri-Mon/Tues). • Get cells, make observation (notebook): color of media, confluency, shape of cells etc.

  10. Second Day (Tuesday, Jan 22) • Check cells. • Check sterility of media. • Decide on and take action.

  11. When ready to passage/split CHO cells • Get hood ready • Warm media • Follow protocol on page 9 • Make appropriate notes in notebook

More Related