The effects of freeze/thawing on human synovial fluid observed by 500 MHz 1H magnetic resonance spectroscopy.

OBJECTIVE

To investigate the effect of freeze/thaw and low temperature storage on the biomolecular profile of human synovial fluid (SF) using high resolution (500 MHz 1H) magnetic resonance spectroscopy (MRS).

METHODS

SF was collected from 12 patients undergoing arthroscopic debridement for treatment of moderate osteoarthritis (OA). Six of the larger samples were divided into 5 parts and treated as follows: the first was analyzed with spin-echo MRS soon after arthroscopy (< 24 h); the 2nd, 3rd, and 4th parts were frozen (-75 degrees C) and thawed for a total of one, 5, and 10 freeze/thaw cycles, respectively, followed by MRS analysis; the 5th part was kept in -75 degrees C storage for > or = 1 year before MRS processing. The 6 smaller samples were divided into 2 parts, the first analyzed shortly after extraction (< or = 24 h), while the 2nd was processed after storage at -75 degrees C for > or = 1 year. Changes in measured metabolite levels were tested for significance using paired t tests.

RESULTS

Freeze-thaw cycling had no statistically significant effect on the relative concentrations of endogenous metabolites measured by MRS, though it did alter individual sample results. Prolonged low temperature storage resulted in a significant drop (p < 0.05) in the signal intensities of glucose (45%), N-acetyl glycoproteins (39%), CH2-chain and CH3-terminal and resonances of lipoproteins (46 and 37%, respectively), valine (43%), leucine (35%), and isoleucine (43%).

CONCLUSION

This study raises questions about routine procedures that may inadvertently affect the outcomes of quantitative SF analyses. Extended low temperature storage should be avoided as it permanently alters the biochemical profile of SF, possibly leading to erroneous conclusions about the nature of OA related changes in metabolite levels with disease progression.