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Phagocytosis VIEW PRODUCT LISTING ASK A QUESTION
- Vial of Monocytes (catalog # 1008 or 1009)
- Alexa Fluor 488 Phalloidin (or other equivalent)
- Alexa Fluor 594 Labeled Zymosan (or other equivalent)
- PBS (Phosphate Buffered Saline)
- Thaw monocytes and suspend in medium at a concentration of 1 million per mL.
- Place coverslips into a petri dish or other suitable container and pipet 250–500 microliters of the cells onto the coverslip. Surface tension should keep the fluid on the coverslip.
- Prepare zymosan according to manufacturer’s recommendations and count particles to determine concentration.
- Add zymosan to the cells at a 10:1 ratio, 10 yeast to each monocyte.
- Incubate at 37°C for one hour. Incubating a coverslip at 2–8°C will provide a negative control.
- Rinse the coverslips gently with 3–4 exchanges of PBS.
- Fix the cells by adding 2% paraformaldehyde to cover the cells and incubate 10 minutes at ambient temperature.
- To counterstain, add 0.1% Triton X-100 to cover the cells for 5 minutes.
- Rinse three times with PBS.
- Add 200 microliters of diluted Alexa Fluor 488 phalloidin.
- Incubate 20 minutes at ambient temperature.
- Wash three times with PBS.
- Place a small drop of 50% glycerol or other mounting medium on a microscope slide and invert the coverslip, cell side down, onto the slide.
- Examine under a fluorescent microscope.