Test Catageory
فئة التحليل
Hematology
Refrence Books
كتب مراجع
1. Daice & Lewis Praactical Hematology
Skills Required
المهارات المطلوبة
1. Pipetting
2. Counting & Calculations
Video Resources
مصادر فيديو
Principle
The manual red cell count requires adequate dilution of blood in a precise ratio using a suitable diluent, thorough mixing and settling of the dilute sample, proper charging and filling of both sides of the hemocytometer chamber, and meticulous counting of erythrocytes in the corner grid areas under high power microscopy. The total number of red cells per unit volume is calculated by applying the dilution factor and hemocytometer dimensions to the averaged counts. Strict adherence to optimized dilution protocols, proficient sample loading, accurate counting, and application of the dilution correction factor is imperative for reliability. Manual enumeration allows determination of absolute red cell numbers when automated technology is inaccessible, given attention to quality of the hemocytometer, dilution preparation, chamber loading, microscopy technique, and mathematical calculation at each step.
Equipments
1. 10 – 100 ul + 100 – 100 ul Automatic Pipette
2. Small glss tubes
3. cholesterol mono-reagent
4. yellow and blue tips
5. Colorimeter or Spectrophotometer
Sample Type
Serum or plasma is acceptable for the Test
But make sure the Patient is fasting to get accurate results
Procedure
1. Prepare the patient’s finger for blood collection using an alcohol swab in case of collecting finger blood. Allow to air dry and Use a sterile lancet to prick the side of the finger and collect 20μl of blood using a capillary tube or Pipette, “in case of EDTA only collect blood in tube using syringe and mix the sample well”.
2. in clean small tube Add 380μl of glacial acetic acid
3. Add the 20μl blood sample to 380μl of Hyme’s Fluid in a test tube.
| Test (T) | |
|---|---|
| Hyme’s Solution | 380 μl |
| Sample | 20 μl |
Procedure
4. Mix the diluted blood well by gently shaking. Allow to stand for 2-3 minutes.
5. Charge the hemocytometer chamber by placing a cover slip over the platform and adding 10μl of diluted blood sample under the cover slip on each side using a pipette.
6. Let stand for 2-3 minutes to allow cells to settle before counting.
7. Count all RBCs in the 4 outer corner squares under 40X objective lens. Count cells touching the middle line on top and left.
8. Repeat counting on the other side chamber.
9. Take the average count of the 4 squares on both sides (N).
10. Calculate RBCs per microliter using the formula: RBCs/microliter = N x Dilution Factor x 104 (Where dilution factor is 20)
11. Report the RBC count in millions/microliter.
M. Sulieman
mohammad@mlsgaate.com
Mix diluted sample well before charging the chamber.
Allow samples to settle before counting.
Take average of counts on both sides for accuracy.
Handle hemocytometer carefully.
