Large-scale investigation of the olfactory receptor space using a microfluidic microwell array

Abstract

The mammalian olfactory system is able to discriminate among tens of thousands of odorant molecules. In mice, each odorant is sensed by a small subset of the approximately 1,000 odorant receptor (OR) types, with one OR gene expressed by each olfactory sensory neuron (OSN). However, the sum of the large repertoire of OR/OSN types and difficulties with heterologous expression have made it almost impossible to analyze odorant responsiveness across all OR/OSN types. We have developed a microfluidic approach that allowed us to screen over 20,000 single cells at once in microwells. By using calcium imaging, we were able to detect and analyze odorant responses of about 2,900 OSNs simultaneously. Importantly, this technique allows for both the detection of rare responding OSNs as well as the identification of OSN populations broadly responsive to odorants of unrelated structures. This technique is generally applicable for screening large numbers of single cells and should help to characterize rare cell behaviors in fields such as toxicology, pharmacology, and cancer research.

Novel cell-based odorant sensor elements based on insect odorant receptors

“…Recently, development of chemical sensing elements has focused on biological molecules or receptor proteins (Feng et al, 2007;Figueroa et al, 2010;Kim et al, 2009;Saito et al, 2009;Sakata and Miyahara, 2008;Radhika et al, 2007;Vidic et al, 2008). A number of cell-based odorant sensors with high sensitivity have been studied and proposed; however, practical development has been inhibited due to shortcomings unique to cells expressing receptor proteins (Du et al, 2013).…”

Highly sensitive and selective odorant sensor using living cells expressing insect olfactory receptors

“…This reaction means that chemical signals are converted to amplified current signals in living systems; the system can be regarded as a transistor with an excellent amplifier. Thus, living cells represent attractive tools for realizing highly selective and sensitive chemical sensors (10,11).…”

Membrane protein-based biosensors

“…As detectors, the fluorescent-based method requires a fluorescence measurement system such as fluorometric imaging plate reader, whereas the latter method is exemplified by an electroantennogram (EAG) that directly detects the action potential of OSNs in vivo, and cultivation of OSNs on a metal oxide semiconductor have been reported as described below. Using a fluorescent indicator for Ca 2þ in OSNs, four different fruity/floral smells (vanilla, rose, berry and banana) were simultaneously distinguished using the responses of approximately 3000 OSNs isolated from a mouse [96]. The approach was based on the fluorescent analysis of numerous of OSNs, which were arrayed in a microfluidic chamber.…”