Cancer Diagnosis Program | Biorepositories and Biospecimen Research Branch

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Effects of certain pre-analytical factors on the performance of plasma phospho-tau217.

Author(s): Bali D, Hansson O, Janelidze S

Publication: Alzheimers Res Ther, 2024, Vol. 16, Page 31

PubMed ID: 38331843 PubMed Review Paper? No

Purpose of Paper

Conclusion of Paper

Studies

1. Study Purpose

   This study compared p-Tau217 levels in plasma from Aβ42/Aβ40 positive and negative patients that were thawed at room temperature versus on wet ice, centrifuged versus not centrifuged before analysis, and in plasma that was subjected to 1, 2 or 3 freeze-thaw cycles.  Additionally, the authors compared the strength of the correlation between plasma p-tau217 levels and CSF Aβ42/Aβ40 and p-tau217 levels among plasma specimens that were thawed at room temperature or on ice and those that were centrifuged or not centrifuged before analysis. Blood was collected into K2EDTA Vacutainer tubes from 50 patients who had a CSF positive Aβ42/Aβ40 result (35 cognitively unimpaired, 6 with mild cognitive impairment, one with Alzheimer’s dementia and one with vascular dementia) and 50 patients who had a CSF negative Aβ42/Aβ40 result (48 cognitively unimpaired, one with Alzheimer’s dementia and one with vascular dementia) . CSF was obtained from the same patients by lumbar puncture.  Plasma and CSF were obtained by centrifugation at 2000 g for 10 min at 4°C and stored at -80°C within 30 min of collection. To investigate the effects of thaw temperature and post-thaw centrifugation, matched aliquots of each plasma specimen were thawed at room temperature or on ice and left uncentrifuged or centrifuged at 2000 g for 10 min. To investigate the effects of freeze-thaw cycling, matched aliquots of plasma were thawed at room temperature and refrozen 0, 1 or 2 times (1-3 cycles) before a final thaw at room temperature; plasma aliquots were then either centrifuged or not. Levels of p-tau217 and Aβ42/Aβ40 were quantified by immunoassay using a MSD SQ120 plate reader.

   Summary of Findings:

   Plasma p-tau217 levels were higher in specimens that were thawed at room temperature and not centrifuged compared to those that were thawed at room temperature and centrifuged (P<0.001, both groups), thawed on ice and not centrifuged (P<0.001 and P=0.005 in Aβ42/Aβ40 negative and positive patients, respectively), and thawed on ice and centrifuged (P<0.001, both groups). Plasma p-tau217 levels were lower in specimens that were thawed at room temperature and centrifuged than in plasma thawed at room temperature and not centrifuged (P<0.001 and P=0.021 in Aβ42/Aβ40 negative and positive patients, respectively). Plasma p-tau217 levels were higher in specimens thawed on ice and not centrifuged compared to those that were centrifuged, but only in specimens from Aβ42/Aβ40 negative patients (P<0.001).

   Plasma p-tau217 levels were modestly and negatively correlated (R=-0.515 to -0.636, P<0.001) with  CSF Aβ42/Aβ40 levels, regardless of thaw temperature and post-thaw centrifugation, with no significant differences in the strength of the correlation. When stratified by Aβ42/Aβ40 status, plasma p-tau217 levels in the Aβ42/Aβ40 positive group were negatively correlated with CSF Aβ42/Aβ40 levels when plasma specimens were centrifuged after thawing (R=-0.394, P=0.01, for room temperature thaw and R=-0.406, P<0.007 for ice thaw), which was not true for uncentrifuged specimens. Plasma p-tau217 levels in the Aβ42/Aβ40 negative group were not correlated with CSF Aβ42/Aβ40 levels. The AUC for plasma p-tau217 detection of abnormal CSF Aβ42/Aβ40 was high (0.845-0.884, sensitivity 76-88%, and specificity 72-90%). While not significantly different, the AUC for specimens that were thawed at room temperature and not centrifuged was slightly lower than for the other conditions examined (0.845 versus 0.872-0.884).

   p-tau217 levels were modestly to strongly correlated between plasma and CSF specimens (R=0.614-0.717, P<0.001), regardless of thaw temperature and post-thaw centrifugation, with no significant differences in the strength of the correlation. When stratified by Aβ42/Aβ40 status, p-tau217 levels were significantly correlated between plasma and CSF specimens from patients that were Aβ42/Aβ40 positive (R=0.506-0.579, P<0.001), regardless of thaw temperature and post-thaw centrifugation, with no significant differences in the strength of the correlation. However, plasma and CSF p-tau217 levels were only correlated when specimens from Aβ42/Aβ40 negative patients were centrifuged after they were thawed (R=0.394, P=0.007, for room temperature thaw and R=0.334, P=0.022 for ice thaw). The AUC, sensitivity and specificity for plasma p-tau217 detection of abnormal CSF p-tau217 were high (0.866-0.924, 88-95%, and 68-83%, respectively).

   Plasma p-tau217 levels were not affected by up to 3 freeze-thaw cycles, but levels were lower in centrifuged than non-centrifuged specimens regardless of the number of freeze-thaw cycles (P<0.001, all). The AUC, sensitivity and specificity plasma p-tau217 of abnormal CSF p-tau217 levels were high regardless of the number of freeze-thaw cycles (0.845-0.876, 72-86% and 78-94%, respectively).

   Biospecimens

   • Bodily Fluid - Cerebrospinal Fluid
   • Bodily Fluid - Plasma

   Preservative Types

   • Frozen

   Diagnoses:

   • Normal
   • Other diagnoses
   • Alzheimer's Disease

   Platform:

   Analyte	Technology Platform
   Protein	Immunoassay

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Storage	Thaw temperature/condition	On ice Room temperature
   Storage	Freeze/thaw cycling	1 cycle 2 cycles 3 cycles
   Biospecimen Aliquots and Components	Centrifugation	Different number of centrifugation steps compared

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