Maintenance and preparation of C albicans hyphae

1 Maintain C. albicans by weekly transfer on Sabouraud dextrose agar plates at 28°C.

2 Transfer a colony from a three-days-old culture to 5 ml PBS in 15 ml polypropylene tube using a sterile loop.

3 Wash twice with PBS by centrifugation at 200 g for 10 min.

4 Resuspend the yeasts at 4 x 10s/ml in complete RPMI medium,

5 Plate 0.1 ml (5 x lO4 yeasts} per well in 96-weIl, flat-bottom microtitre plates with a repeating dispenser.

6 Incubate plates for 3-6 h at 37 °C until 95% of Candida have germinated into hyphae (30-100 ^m in length) that adhere firmly to the bottom of the wells (check hyphae germination by inverted microscope examination).3

7 Remove supernatant from each well by pipette aspiration observing biohazard precautions.

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