Background: Iron deficiency (ID) may impair long-term neurological development when it occurs in young infants. In cohort studies, it is sometimes necessary to evaluate ID with sera kept frozen for several years. To assess ID, learned societies recommend measuring serum ferritin (SF) level combined with C-reactive protein level. The long-term stability of C-reactive protein in frozen samples is well established but not ferritin.
Methods: We measured SF level (immunoturbidimetric assay; in micrograms per liter) immediately after collection from 53 young adults recruited and followed-up in Porto, Portugal, from 2011 to 2013 (SF1), and then, in 2016 in two aliquots kept frozen at- 80°C for 3 to 5 years: one without (SF2A) and one with (SF2B) intermediate thawing in 2014. We compared SF1 to SF2A then SF2B; statistical agreement was evaluated by the Bland and Altman method and the effect of intermediate thawing by regression modelling.
Results: Mean SF2A-SF1 and SF2B-SF1 differences were -2.1 (SD 7.0) and 48.9 (SD 66.9). Values for Bland and Altman 95% limits of agreement were higher for the comparison of SF2B and SF1 than SF2A and SF1: -82.2 to 179.9 and -15.8 to 11.8, respectively; the effect of thawing was highly significant (p <0.001).
Conclusions: Agreement between SF values before and after 3 to 5 years of constant freezing at -80°C was in a generally accepted range, which supports the hypothesis of ferritin's stability at this temperature for a long period. In long-term storage by freezing, intermediate thawing induced a major increase in values.