Immunochemical Quantitation of Proteins in Single Cells

T HE NATURE of the transition from hemoglobin F synthesis to hemoglobin A synthesis in the fetus and newborn remains unknown. Although it is evident from the elution method developed by Kleihauer, Braun and Betke13 that hemoglobin F and hemoglobin A may coexist in single erythrocytes,24 quantitative data regarding the amounts of these hemoglobins in individual cells are lacking. An attempt to quantitate specific hemoglobins in single erythrocytes by combining microspectrophotometry with the elution technic has not proven very successful, the method being insensitive to levels of hemoglobin F which were less than 10 per cent or more than 80 per cent of the total.5 Excellent methods for the quantitation of total hemoglobin in single cells have been developed using photometry6 or electron microscopy,7 but the methods do not distinguish between specific hemoglobins. Immunochemical methods have been used successfully to differentiate between cells which do or do not contain hemoglobin F,81#{176} but the methods are qualitative, not quantitative. In the present study, specific antisera against y-chain hemoglobin, /3-chain hemoglobin and carbonic anhydrase B were used to determine the quantities of these proteins in single erythrocytes in normal fetal, newborn and adult blood. In addition, cells from patients with sickle cell hemoglobin or with thalassemia were examined for y-chain hemoglobin and carbonic anhydrase B.