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Homogeneous assay for whole blood folate using photon upconversion
Riikka Arppe1, Leena Mattsson, Krista Korpi
1Department of Biochemistry/Biotechnology, University of Turku , Tykistökatu 6 A, 20520 Turku, Finland.
Analytical Chemistry
|December 31, 2014
Summary
This study introduces a novel photon upconversion assay for measuring red blood cell folate directly in whole blood. This method overcomes limitations of traditional assays, offering a simpler and faster diagnostic tool for folate deficiency.
Area of Science:
- Biomedical Diagnostics
- Nanotechnology in Medicine
- Analytical Chemistry
Background:
- Red blood cell folate is a key indicator for diagnosing folate deficiency, reflecting long-term tissue levels.
- Conventional fluorescence assays for whole blood folate face challenges due to high background and light absorption.
- Existing methods often rely on serum folate, which only indicates recent dietary intake, not sustained levels.
Purpose of the Study:
- To develop a novel, direct, homogeneous assay for measuring red blood cell folate in whole blood.
- To overcome the limitations of conventional fluorescence techniques in complex biological samples like blood.
- To utilize photon upconversion and resonance energy transfer for enhanced sensitivity and reduced interference.
Main Methods:
- A photon upconversion-based homogeneous assay was developed using upconverting nanophosphors coated with folate binding protein.
- Resonance energy transfer was employed between the nanophosphor donor and a near-infrared fluorescent acceptor dye conjugated to a folate analogue.
- Measurements were taken at 740 nm upon 980 nm excitation, minimizing sample absorption and autofluorescence via anti-Stokes detection.
Main Results:
- The assay demonstrated a low limit of detection (LOD) of 1 nM and an IC50 value of 6.0 nM.
- A measurable concentration range of two orders of magnitude (1.0-100 nM) was achieved, corresponding to 40-4000 nM in whole blood.
- High recoveries (112%-114%) for added folic acid and good correlation with a DELFIA-based assay were observed.
Conclusions:
- The developed photon upconversion assay offers a simple, fast, and sensitive method for direct whole blood folate measurement.
- This technique effectively minimizes background noise and light absorption issues inherent in conventional blood assays.
- The assay holds promise for improved diagnosis of folate deficiency by providing a more accurate reflection of folate status.
