Staff Publications

Staff Publications

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    'Staff publications' is the digital repository of Wageningen University & Research

    'Staff publications' contains references to publications authored by Wageningen University staff from 1976 onward.

    Publications authored by the staff of the Research Institutes are available from 1995 onwards.

    Full text documents are added when available. The database is updated daily and currently holds about 240,000 items, of which 72,000 in open access.

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Record number 553498
Title The Effect of Calcium Buffering and Calcium Sensor Type on the Sensitivity of an Array-Based Bitter Receptor Screening Assay.
Author(s) Roelse, M.; Wehrens, H.R.M.J.; Henquet, M.G.L.; Witkamp, R.F.; Hall, R.D.; Jongsma, M.A.
Source Chemical Senses 44 (2019)7. - ISSN 0379-864X - p. 497 - 505.
DOI https://doi.org/10.1093/chemse/bjz044
Department(s) BIOS Applied Metabolic Systems
Biometris
Mathematical and Statistical Methods - Biometris
Nutritional Biology and Health
VLAG
Bioscience
Laboratory of Plant Physiology
Publication type Refereed Article in a scientific journal
Publication year 2019
Abstract The genetically encoded calcium sensor protein Cameleon YC3.6 has previously been applied for functional G protein–coupled receptor screening using receptor cell arrays. However, different types of sensors are available, with a wide range in [Ca2+] sensitivity, Hill coefficients, calcium binding domains, and fluorophores, which could potentially improve the performance of the assay. Here, we compared the responses of 3 structurally different calcium sensor proteins (Cameleon YC3.6, Nano140, and Twitch2B) simultaneously, on a single chip, at different cytosolic expression levels and in combination with 2 different bitter receptors, TAS2R8 and TAS2R14. Sensor concentrations were modified by varying the amount of calcium sensor DNA that was printed on the DNA arrays prior to reverse transfection. We found that ~2-fold lower concentrations of calcium sensor protein, by transfecting 4 times less sensor-coding DNA, resulted in more sensitive bitter responses. The best results were obtained with Twitch2B, where, relative to YC3.6 at the default DNA concentration, a 4-fold lower DNA concentration increased sensitivity 60-fold and signal strength 5- to 10-fold. Next, we compared the performance of YC3.6 and Twitch2B against an array with 11 different bitter taste receptors. We observed a 2- to 8-fold increase in sensitivity using Twitch2B compared with YC3.6. The bitter receptor arrays contained 300 spots and could be exposed to a series of 18 injections within 1 h resulting in 5400 measurements. These optimized sensor conditions provide a basis for enhancing receptomics calcium assays for receptors with poor Ca2+ signaling and will benefit future high-throughput receptomics experiments.
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