CONTINUOUS MEASUREMENT OF THE CHANGE IN ERYTHROCYTE VOLUME AFTER ADDITION OF SAPONIN (Saponin Test)
One μ1 of whole blood is added to 50 ml aliquots of 0.9 % and 0.5% saline solutions prewarmed at 30°C and mixed to get homogeneous suspensions of erythrocytes. Into each of these suspensions two hundred μ1 of 1% aqueous solution of saponin is put 40 seconds after addition of blood to the saline solutions. The changes in mean erythrocyte volume (MCV) are able to observe continuously in an automatic blood cell counter equipped with a recording assembly. The results of the observations are recorded in graphic curves. The curves analyzed for the expansion ratio (E), the time (in sec.) required for attaining the maximum MCV (T max), and the time (in sec.) necessary for erythrocytes to reach the critical point of complete rupture due to erosive effect exerted by saponin (T end). This observation or test is called Saponin test. Saponin test of erythrocytes was carried out with heparinized blood samples collected from normal subjects. The results obtained showed that initial mean corpuscular volume (IMCV) was 85-91 fl, maximum value of mean corpuscular volume (MMCV) 108-118 fl, E 128-137%, T max 28-33 sec., T end (in 0.9% saline) 38-44 sec. and T end (in 0.5% saline) 30 57 sec. The test was applied to the examination of physicochemical properties of erythrocytes of the patients with hereditary spherocytosis (HS), β-thalassemia (β-Thal), iron deficiency anemia (IDA), diabetes mellitus (DM) and hepatobiliary disorders. The test results were examined in comparison with the lipid compositions of relevant erythrocyte membranes which were analyzed simultaneously. β-Thal gave a pathognomonic pattern of saponin test. (Decrease in IMCY and MMCY, increase in E, and prolongation of T max and T end). IDA was discriminated from β-thalassemia by normal or slightly shortened T max and T end. HS was characterized by remarkable decrease in E. The erythrocytes of hepatobiliary disorders in which increase in free cholesterol of the cellular membrane was commonly evident were very sensitive to saponin test. Their T max and T end were shortened. The abnormality in saponin test was not significantly remarkable in DM. The saponin test is thought to be a new test recommended for the differential diagnosis of β-thalassemia from iron deficiency anemia.