Physical examination See for the following physical characteristics

• Volume
• Colour
• Appearance

Microscopic Examination Cell count should be made soon after collection of fluid.Leukocyte cell count is done with the help of Neubaur’s chamber and micropipette.

Method

1. Mix the fluid well.
2. Prepare direct examination of fluid by putting one drop over a slide and lay a coverslip over it.
3. Take 100 microlitres of fluid into one test tube and mix it with 100 microlitres of WBC diluting fluid.Mix it and keep for 10-15 minutes.
4. Mixed well and Neubaur’s chamber is charged as for WBC Count.
5. The rulings are brought into focus by using low power objective.The cells in the four larger corner squares of Neubaur’s chamber are counted.
6. Perform microscopic examination as follows.

WBC DILUTING FLUID: The diluting fluid used should stain the leukocytes distinctly and lyse RBC’s.

Most commonly used is

• 10% Glacial acetic acid………………………….2.0 ml
• 1%(w/v) Gentian violet………………………….1.0 ml
• Distilled water……………………………………97 ml.

Calculation Total Leukocyte count :

                      N     X    D
                    ______________
                            V                            
                                     
                     : N   X   2  
                     ______________   	
                        1X1X0.1 
                        
                     :  N X20/ CU. MM

N: average number of leukocyte counted in one square D: Dilution V: Volume of 1 large square

Differential count of cells Sample is centrifuged at 3000-3500 rpm for 5 min. Examine color of supernatant and smear is prepared from the centrifuge deposit, dried and stained by Giemsa stain. Differential leukocyte count is done by counting 500 cells manually on giemsa stain sediment smear. The smear is cross checked by senior person simultaneously.