acid hemolysis test

Summary

Erythrocytes from patients with PNH, due to their inherent defects, have increased sensitivity to complement and are easily destroyed by hemolysis in acidified normal serum, (PH 6.0-6.8) by incubation at 37°C. This method is more sensitive and has fewer false positives. This experiment is from the Mudanjiang Medical College undergraduate 5-year laboratory guide for testing majors

Operation method

acid hemolysis test

Principle

Erythrocytes from patients with PNH, due to their inherent defects, have increased sensitivity to complement and are easily destroyed by hemolysis in acidified normal serum, (PH 6.0-6.8) by incubation at 37°C. This method is more sensitive and has fewer false positives.

Materials and Instruments

Peripheral blood
HCl NaCI solution
Centrifuge tube

Move

I. Experimental reagents:

1. O.2mol/L HCI

2. 8.5g/L NaCI solution

II. Experimental manipulation:

l. Use 10ml centrifuge tube 1, add 8.5g / L NaCI solution 5-6ml, then add more than 10 drops of the patient's peripheral blood. Mix well and centrifuge to remove the supernatant. Repeat the washing of erythrocytes for 2 times, and finally formulate 50% erythrocyte suspension.

2. Take the same type as the patient or AB blood 3-5ml, to be its natural coagulation, separation, normal human serum (preferably mixed with a number of fresh serum to ensure that there is enough complement).

3. Prepare 50% erythrocyte suspension from the same normal human blood as above according to operation "l", and operate according to the table as a normal control.

4. Place in a 37℃ water bath for 1 hour and observe the results.

5. Judge the results of the experiment:

(1) Test tube hemolysis control tube does not hemolysis is positive.

(2) Negative if neither the test tube nor the control tube is hemolyzed.

Caveat

1. After acidification of the serum. The test tube must be plugged tightly, otherwise carbon dioxide will escape. Otherwise, carbon dioxide can escape and decrease the acidity of the serum.

2. Anticoagulants will affect the pH value, so it is not suitable to use anticoagulant serum． But can be used to defibrillate blood.

3. In order to ensure adequate complement, it is best to use mixed serum, but the normal control red blood cells must be used "O" type.

4. If the patient has been transfused several times, the abnormal red blood cells in the blood are relatively reduced. The test may be weakly positive or negative.

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