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Two hundred and twenty-three subjects out of a total of 347 with various degrees of glucose tolerance were recalled after a screening survey for diabetes. They were a randomly selected sample of people over the age of 40 and they underwent a formal 75 g glucose tolerance test in order to assess the effect of a glucose load on glycohaemoglobin levels measured by four different assay methods.Oral glucose loading was found to affect glycohaemoglobin levels only when these were measured by an agar-gel electrophoretic method that did not remove the labile aldimine-linked Schiff base fraction. The increase in glycohaemoglobin during the glucose tolerance test as estimated by this method was proportional to the 2 h blood glucose level. Glycohaemoglobin levels measured by agar-gel electrophoresis with elimination of the Schiff base, by affinity chromatography and by iso-electric focussing, were not affected by a 75 g oral glucose load. We conclude that blood samples for glycohaemoglobin assay may be collected at any time of the day, without regard to the subject's previous food intake, provided an assay method is used that removed the aldimine-linked labile fraction.