Automated assays and methods for detecting and modulating cell surface protein function

An automated drug screening assay for identifying compounds that modulate activity of ion channels and/or receptors of a cell, comprising

• (a) introducing a divided culture vessel having an array of individual compartments into an apparatus, wherein:
• at least two compartments contain cells to be assayed comprising ion channels and/or receptors that when activated, directly or indirectly, cause a detectable change in the concentration of a predetermined ion in the cytoplasm of the cells;
• the cytoplasm of the cells comprises an amount of an ion-sensitive fluorescent indicator sufficient to detect a change in the concentration of the predetermined ion;
• and
• the apparatus delivers reagent solution to one or more compartments of the divided culture vessel and detects and measures an attribute of the contents of the individual compartments;
• (b) automatically delivering to one or more predetermined compartments an aliquot of a solution comprising either or both of (i) an amount of a known ion channel- or receptor-activating compound that is effective to activate the ion channel or receptor, and (ii) a test compound of unknown ion channel or receptor activity;
• and
• (c) within a predetermined time that is less than an amount of time for any induced ion flux to reach a maximum, detecting and measuring, in one or more of the individual compartments for a predetermined amount of time, the induced ion flux by detecting or measuring fluorescence emitted by the ion-sensitive indicator in response to an excitation wavelength, whereby compounds that modulate activity of ion channels and/or receptors of a cell are identified.

An automated method for detecting or measuring an occurrence of a cellular event that directly or indirectly induces a change in intensity of an optical attribute of an indicator moiety in response to the cellular event, comprising:

• (a) introducing a divided culture vessel having an array of individual compartments into an apparatus, wherein:
• at least one compartment contains cells that, when contacted with a compound that causes the cellular event to occur, undergo the cellular event;
• the cytoplasm of the cells comprises an amount of the indicator moiety sufficient to exhibit a detectable change in the intensity of the optical attribute upon occurrence of the event;
• and
• the apparatus has delivery means to deliver reagent solution to one or more compartments of the divided culture vessel and means to detect and measure at least one optical attribute of the contents of the compartments;
• (b) automatically delivering to one or more predetermined compartments, an aliquot of a solution comprising an amount of a compound that causes the cellular event to occur thereby allowing a detectable increase in the intensity of the optical attribute in the compartment containing the cells to be measured, wherein the delivery means and predetermined compartments are aligned;
• and
• (c) within a predetermined time period that is less than the time for the cellular event indicated by the change in the intensity of the optical attribute to reach its peak, detecting and measuring in one or more of the predetermined compartments for a predetermined amount of time the intensity of the optical attribute emitted by the cells, wherein the detecting and measuring means and predetermined compartments are aligned, and whereby the occurrence of a cellular event is detected or measured.

An automated method for identifying compounds that induce the occurrence of a cellular event that results in a change in the intensity of an optical attribute of an indicator moiety in response to the cellular event, the method comprising:

• (a) introducing a divided culture vessel having an array of individual compartments into an apparatus, wherein:
• (i) at least one compartment contains cells that when contacted with a compound that causes the cellular event to occur, undergo the cellular event;
• (ii) the cytoplasm of the cells comprises an amount of an indicator moiety sufficient to exhibit a detectable change in the intensity of the optical attribute upon occurrence of the event;
• and
• (iii) the apparatus has delivery means to deliver reagent solution to one or more compartments of the divided culture vessel, and detecting and measuring means to detect and measure at least one attribute of the contents of the compartments;
• (b) automatically delivering to one or more predetermined compartments an aliquot of a solution comprising either or both of (i) an amount of a known compound that is effective to induce the cellular event;
• and (ii) a test compound of unknown activity;
• and
• (c) within a predetermined time period that is less than the time for the cellular event indicated by the change in the intensity of the optical attribute to reach a maximum, detecting and measuring in one or more of the predetermined compartments, for a predetermined amount of time the level of the optical attribute emitted by the cells, wherein the detecting and measuring means and the predetermined compartments are aligned, whereby a compound that induces the occurrence of a cellular event is identified.

A method for automatically measuring an optical attribute indicative of occurrence of a transient reaction in a computer-controlled measurement apparatus, the method comprising:

• (a) introducing an ion-sensitive indicator into cells in a plurality of wells of a multi-well plate containing samples to be measured;
• (b) identifying at least one predetermined well of a plurality of wells of a multi-well plate containing samples to be measured;
• (c) aligning the predetermined well(s) by the apparatus with a predetermined position;
• (d) adding reagent by the apparatus to the predetermined well(s) at the predetermined position(s), wherein the reagent induces directly or indirectly the transient reaction;
• (e) measuring the optical attribute of the predetermined well(s) while aligned with the position(s), wherein commencement of measurement occurs within a predetermined time period that is less than the time required for the reaction to reach its peak response and measurement is effected for a predetermined amount of time, whereby the optical attribute indicative of a transient reaction is automatically measured in the computer-controlled apparatus.