Subculture of a cell monolayer

Take some cell suspension and dilute with an equal volume of 0.1% trypan blue (stains dead cells Appendix 2). 

Take up some trypan blue cell suspension in a Pasteur pipette and fill a haemocytometer counting chamber by capillary attraction ( 7.2.1). Take care not to flood the channels of the chamber. Count the cells under X10 objective. 

Count the four sets of 16 small squares (on each corner of the central ruled area ( 7.2.1). Count only unstained cells. Divide by 4 to give the average count per 1 mm2. Since the area is 1 mm2 and the depth 0.1 mm, the conversion factor for the counting chamber is 104, e.g. let the average count = 20 cells per 1 mm2 

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