Fig. 1

Schematic representation of pre-analytical sample handling conditions. a The blood was collected in 6 EDTA tubes from each of the 100 study participants and centrifuged at 2000 g at 4 °C for 10 min. After centrifugation, plasma was transferred from all six EDTA tubes into one 50-ml tube, mixed, and 1 ml was aliquoted into 1.5-ml tubes and stored at − 80°C within 30 min after collection. For each participant, two tubes with 1 ml of frozen plasma were used. One frozen plasma tube was thawed on ice while the other tube was thawed at RT. Following thawing, 200-μl plasma aliquots were prepared. For samples thawed at RT, one 200-μl plasma aliquot was centrifuged at RT (10 min, 2000 g) whereas another 200-μl plasma aliquot was not centrifuged and stored at RT until the p-tau217 analysis. The plasma aliquots prepared from the tube thawed on ice were treated in the same way except that centrifugation and sample storage prior to the p-tau217 analysis were performed at + 4 °C and on ice respectively. b To assess the effect of freeze–thaw cycles, a new 1-ml plasma tube was used for each participant. The frozen plasma tube was thawed at RT and four 70-μl plasma aliquots were prepared. Two of the four plasma aliquots did not undergo any further freeze–thaw cycles and were thus fxt-1; the other two 70-μl plasma aliquots underwent one additional freeze–thaw cycle (fxt-2) within 30 min prior to analysis. Additionally, two plasma aliquots that were fxt-3 times (prepared from the previous set of experiments) were also included. One aliquot of each of the three freeze–thaw conditions (fxt-1, 2, or 3) was centrifuged at RT whereas another aliquot was not centrifuged and stored at RT until the p-tau217 analysis. Abbreviations: Cond, Condition; EDTA, ethylenediaminetetraacetic acid; fxt, freeze–thaw cycle; RT, room temperature. Created with BioRender.com