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There is no general agreement regarding the use of the first or second drop of blood for glucose monitoring. This study investigated whether capillary glucose concentrations, as measured in the first and second drops of blood, differed ≥10% compared with a control glucose concentration in different situations.Capillary glucose concentrations were measured in two consecutive drops of blood in the following circumstances in 123 patients with diabetes: without washing hands, after exposing the hands to fruit, after washing the fruit-exposed hands, and during application of different amounts of external pressure around the finger. The results were compared with control measurements.Not washing hands led to a difference in glucose concentration of ≥10% in the first and in the second drops of blood in 11% and 4% of the participants, respectively. In fruit-exposed fingers, these differences were found in 88% and 11% of the participants, respectively. Different external pressures led to ≥10% differences in glucose concentrations in 5–13% of the participants.We recommend washing the hands with soap and water, drying them, and using the first drop of blood for self-monitoring of blood glucose. If washing hands is not possible, and they are not visibly soiled or exposed to a sugar-containing product, it is acceptable to use the second drop of blood after wiping away the first drop. External pressure may lead to unreliable readings.