	US 12,105,122 B2
	Voltage indicators
Eric R. Schreiter, Ashburn, VA (US); and Ahmed Abdelfattah, Ashburn, VA (US)
Assigned to HOWARD HUGHES MEDICAL INSTITUTE, Chevy Chase, MD (US)
Filed by HOWARD HUGHES MEDICAL INSTITUTE, Chevy Chase, MD (US)
Filed on Jan. 27, 2023, as Appl. No. 18/160,764.
Application 18/160,764 is a continuation of application No. 16/872,188, filed on May 11, 2020, granted, now 11,598,792.
Claims priority of provisional application 62/845,643, filed on May 9, 2019.
Prior Publication US 2023/0296649 A1, Sep. 21, 2023
Int. Cl. G01R 15/22 (2006.01); C07K 14/195 (2006.01); C07K 14/435 (2006.01); G01N 21/64 (2006.01); G01R 19/00 (2006.01)

CPC G01R 15/22 (2013.01) [G01R 19/0084 (2013.01); C07K 14/195 (2013.01); C07K 14/43504 (2013.01); C07K 2319/60 (2013.01); G01N 21/6486 (2013.01)]

20 Claims

1. A method of measuring voltage, the method comprising

(a) providing a voltage indicator, comprising:

(i) a voltage-sensitive microbial rhodopsin domain comprising a polypeptide selected from a group consisting of QuarsAr1, QuarsAr2, Ace2N, and combinations thereof including one, two, three, or four amino acid mutations relative to a wild type polypeptide sequence; and

(ii) a capture protein that covalently or noncovalently binds a fluorescent dye ligand that is a fluorescent protein; or

a fluorescent dye, wherein the capture protein is provided together with the voltage-sensitive microbial rhodopsin domain in a fusion protein-;

(b) contacting the voltage indicator and the fluorescent dye ligand with a cell, and

(c) determining changes in fluorescence of the fluorescent dye ligand when the fluorescent dye ligand is captured by the voltage indicator.