Erythrocyte counting assay

Summary

Knowledge of the principles and techniques of erythrocyte counting (microscopic imaging)

Operation method

Erythrocyte counting assay

Principle

Blood is diluted a certain number of times with isotonic diluent, filled into the counting ground, and the number of red blood cells in a certain volume is counted under the microscope, and the number of red blood cells per liter of blood is converted to show the number of red blood cells in the blood.

Materials and Instruments

Hermitage Red Diluent Sodium Chloride Crystallized Sodium Sulfate High Mercury Chloride Distilled Water Lemon Salt Methylated Salt Water
Microscope, hemoglobin pipette, counting plate, coverslip.

Move

III. Experimental reagents:

l, Hayem (Hayem) generation of red fine ground dilution solution

Sodium chloride (NaCl) 0.1g

Crystallized sodium sulfate 5.0g

(or Na2SO4) 2.5g

mercuric chloride (HgC12) 0.5g

Distilled water Add to 20Oml and filter and use the sodium chloride in it. Sodium sulfate acts as an osmotic concentration regulator, and the other Na2SO4 also prevents red blood cell aggregation; HgCI2 is a preservative.

2, lemon into the salt a stay brine

NaCl 0.6g

Sodium citrate (Na3C6H5O7?2H2O) 1.0g

Formalin 1mL

Distilled water Add to 100mL filter and adjust osmotic concentration using NaCl, Na3C6H5O7?2H2O, the latter being anticoagulant and formaldehyde preservative.

3、Newly prepared isotonic saline

IV. Experimental equipment: microscope, hemoglobin pipette, counting plate, coverslip V. Experimental operation:

l, take 1 test tube, add diluent 3.98mL (or 1.99mL)

2, with a clean and dry micropipette accurately suck 10 microliters of peripheral blood.

3, wipe off the tip of the tube outside the remaining blood, gently injected into the bottom of the red blood cell diluent. Re-absorb the upper layer of diluent clear pipette 2-3 times, immediately shake well.

4、Wipe the counting cell and coverslip with soft fabric, cover the coverslip on the counting cell.

5、Using a pipette to suck up the mixed red blood cell suspension, filling the counting pool.

6、Stand still for 2-3 minutes, to be via the erythrocyte sinking, with high or low magnification microscope to count the number of erythrocytes in the center of the four corners of the large square and the middle of the five in the middle of the square. Six, calculation:

N (the number of RBCs in the five middle squares) × 5 × 10 × ¹⁰⁶ × 200 = N × ^1012/liter × 5: indicates that the five middle squares in the RBCovt converted to 1 large square in the number of RBCs × 10: l large square volume, 0.1ul, converted to lul in the number of RBCs

^×106: 1uL is converted to 1 liter of RBCs.

×20O: number of dilutions

VII. Experimental report mode: ^{ΔΔΔ×1012/L}

Normal reference value: adult male: 4.OO-5. ^50×1012/L

Adult female: 3.50-5. ^00×1012/L

Neonate: 6.00-7. ^00×1012/L

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