Microdroplet test apparatus

• US 4,599,315 A
• Filed: 09/13/1983
• Issued: 07/08/1986
• Est. Priority Date: 09/13/1983
• Status: Expired due to Fees

• Current Assignee: Regents of the University of California (University of California)
• Original Assignee: Regents of the University of California (University of California)
• Inventors: Terasaki, Paul I., Aoki, Dennis
• Primary Examiner(s): Warden, Robert J.
• Assistant Examiner(s): KRAWCZEWICZ MYERS, LOUANNE C
• Application Number:
  US06/531,923
• Time in Patent Office: 1,029 Days
• Field of Search: 435/33, 435/293, 435/300, 435/292, 435/294, 435/298, 435/299, 435/301, 435/810, 436/809, 436/180, 422/102, 422/73, 422/99, 356/246, 356/244, 350/536
• US Class Current: 435/288.4
• CPC Class Codes: B01L 3/5085 for multiple samples, e.g. microtitration plates
• CPC Class Codes: C12Q 1/20 using multifield media
• CPC Class Codes: G01N 2035/1034 Transferring microquantities of liquid
• CPC Class Codes: G01N 35/028 having reaction cells in the form of microtitration plates
• CPC Class Codes: Y10S 435/81 Packaged device or kit
• CPC Class Codes: Y10S 436/809 Multifield plates or multicontainer arrays
• CPC Class Codes: Y10T 436/2575 Volumetric liquid transfer

Abstract

A microdroplet test apparatus for use in HLA typing. The apparatus is an improvement upon the well known Terasaki tray which is in use for determining HLA antigens by measuring lymphocyte cytotoxicity. The apparatus includes a tray having a plurality of microtest wells wherein the sides of the microtest wells are designed to promote localization of lymphocytes or other cells being tested at the microtest well bottom. The apparatus further includes a cover having either rods or tubes which extend down into the test solution present in the microtest well to thereby control the amount of test solution which is present in the cylindrical optical view path through which the test solution is viewed by microscope for test result measurements. This allows control of the amount of test solution in the optical view path in order to reduce the amount of solution which must be looked through when cells are being counted or alternatively, increasing the amount of test solution present in the view path in order to maximize detection when a colored reaction product in the test solution is being measured.

Claims

1. A microdroplet test apparatus adapted for use in determining HLA antigens by measuring lymphocyte cytotoxicity, said apparatus comprising:

a tray having a top surface, a bottom surface and a perimeter;

a plurality of microtest wells at spaced locations in said tray for receiving and holding test solutions including lymphocytes and selective cytotoxic reagents capable of lysing selected lymphocytes, said microtest wells including an upper rim on said tray top surface defining an opening in said tray top surface having a surface area sufficient to allow introduction of lymphocytes and reagents into said wells, a bottom having a surface area less than the surface area of said well opening and a well side wall extending between said upper rim and said well bottom, said microtest well including a central cylindrical zone defining a vertical view path wherein lymphocyte cytotoxicity is measured by determining the degree of lymphocyte lysis by viewing vertically down through the test solution present in said vertical view path and wherein said well side wall has a circular rim and the vertical cross section of said wall is arcuate, such that the slope of said side wall is less than vertical adjacent said circular rim and increases to substantially vertical slope adjacent said well bottom to thereby promote localization of said lymphocytes toward said well bottom;

a cover for said tray having a top surface, a bottom surface and a perimeter;

means associted with the perimeters of said tray and said cover for providing a seal therebetween when said cover is placed on said tray;

view path control means located on said cover extending downward from said cover bottom and insertable within said microtest wells for controlling the amount of test solution present in said vertical view path, wherein said view path control means includes a tube attached to said cover and positioned to extend vertically downward into said microtest well when said cover is in position on said tray, said tube having an internal conduit along said view path with an open bottom end which is insertable into said test solution wherein said tube and internal conduit are sized such that said test solution is drawn up into said conduit by way of capillary action to provide an increase in the depth of said test solution in said view path.

2. A microdroplet test apparatus according to claim 1 wherein said means for sealing said cover to said tray is provided by one or more mating tongue and groove surfaces extending around the perimeter of said cover and said tray.

3. A cover for a microdroplet test tray wherein said test tray includes a top surface, a bottom surface, a perimeter and a plurality of microtest wells at spaced locations in said tray for receiving and holding test solutions including lymphocytes and selective cytotoxic reagents capable of lysing selected lymphocytes, said microtest wells including a central vertical cylindrical zone defining a vertical view path, said cover comprising:

a cover plate having a top surface, a bottom surface and perimeter for sealing engagement with the perimeter of said tray;

means associated with said cover plate perimeter for sealing said cover plate to said tray when said cover plate is positioned on said cover plate; and

view path control means located on said cover plate extending downward from said cover plate bottom and insertable within said microtest wells for controlling the amount of test solution present in said vertical view path to thereby provide selective control of the depth of solution in said vertical view path, said view path control means including a tube attached to said cover plate and positioned to extend vertically downward into said microtest well when said cover plate is in position on said tray, said tube having an internal conduit along said view path with an open bottom end which is insertable into said test solution wherein said tube and internal conduit are sized such that said test solution is drawn up into said conduit by way of capillary action to provide said increase in the depth of said test solutions in said view path.

4. In a microtest tray having a top surface, a bottom surface, a perimeter and a plurality of microtest wells at spaced locations in said tray for receiving and holding test solutions including lymphocytes and selective cytotoxic reagents capable of lysing selected lymphocytes, said microtest wells including an upper rim on said tray top surface having a surface area sufficient to allow introduction of lymphocytes and reagents into said wells, a bottom having a surface area less than the surface area of said well opening and a well side wall extending at a constant slope downward between said upper rim and said well bottom, wherein the improvement comprises:

an arcuate side wall having a slope which gradually increases from less than vertical near the well rim to substantially vertical at the well bottom to thereby promote localization of said lymphocytes toward said well bottom.