June 2022
Volume 63, Issue 7
Open Access
ARVO Annual Meeting Abstract  |   June 2022
Effect of Sample Collection and Storage on Biological Stability of Cytokines in Human Aqueous Humor and Vitreous Samples
Author Affiliations & Notes
  • Tina Felfeli
    University of Toronto Institute of Health Policy Management and Evaluation, Toronto, Ontario, Canada
    University of Toronto Department of Ophthalmology and Vision Sciences, Toronto, Ontario, Canada
  • Bret Nestor
    University of Toronto Department of Computer Science, Toronto, Ontario, Canada
  • Jeff Park
    University of Toronto Temerty Faculty of Medicine, Toronto, Ontario, Canada
  • David T Wong
    University of Toronto Department of Ophthalmology and Vision Sciences, Toronto, Ontario, Canada
    Ophthalmology, Unity Health Toronto, Toronto, Ontario, Canada
  • Footnotes
    Commercial Relationships   Tina Felfeli None; Bret Nestor None; Jeff Park None; David Wong None
  • Footnotes
    Support  None
Investigative Ophthalmology & Visual Science June 2022, Vol.63, 2370 – A0054. doi:
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      Tina Felfeli, Bret Nestor, Jeff Park, David T Wong; Effect of Sample Collection and Storage on Biological Stability of Cytokines in Human Aqueous Humor and Vitreous Samples. Invest. Ophthalmol. Vis. Sci. 2022;63(7):2370 – A0054.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose : Cytokines are inherently prone to proteolytic degradation and thus conclusions drawn from collection and storing of the ocular samples could potentially result in misled clinical decisions. This study aimed to determine the effects of sample collection and storage duration on the levels of various cytokines in the human aqueous humor and vitreous samples.

Methods : Samples were obtained from 27 human eyes that underwent pars plana vitrectomy for various diagnoses. Undiluted samples (0.1-0.3ml) were aliquoted into 3 tubes and stored at -80oC for analysis within 1-week, 3-month and 9-month duration since collection. All 27 cytokine analytes were analyzed with the Bioplex Pro-Human cytokine 27-plex assay kit. An ANOVA was run to assess the impact of storage time and sampling location for each biomarker. Cytokine profiles were embedded using principal component analysis (PCA) to view the proximity between timepoints for each sample. A p-value less than the Bonferroni corrected threshold of 0.05 was considered significant.

Results : Four of the 27 biomarkers were significantly impacted by storage duration at both 3- and 9-month timepoints (Interleukin 2 [IL-2], IL-10, IL-12, and Platelet-derived growth factor [PDGF-BB]), whereas 11 were significantly influenced by sampling location of anterior chamber versus vitreous within the same eyes (Granulocyte-macrophage colony-stimulating factor [GM-CSF], IL-2, IL-5, IL-6, IL-7, IL-8, IL-9, IL-12, IL-15, PDGF-BB, and vascular endothelial growth factor [VEGF]). Amongst biomarkers where sample duration was significant, the relative abundance tended to decrease with time. For biomarkers where sampling location of anterior chamber versus vitreous was significant, cytokine concentrations tended to be higher in aqueous humor, with the exception of IL-7. Neither sampling location offered a significant advantage of mitigating deterioration in storage. Separability of patient-specific cytokine profiles at all 3 timepoints in the PCA remained relatively the same over time.

Conclusions : Although there is degradation amongst specific cytokine analytes over 9 months of sample storage, sampling between aqueous humour and vitreous specimens within the same eyes shows much more variation than individual sample deterioration. In all cases, the overall patient-specific cytokine profiles remained relatively the same over time.

This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.

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