System and method for sorting cells

• US 9,422,523 B2
• Filed: 02/11/2013
• Issued: 08/23/2016
• Est. Priority Date: 12/31/1997
• Status: Expired due to Fees

• Current Assignee: XY, LLC (Inguran LLC)
• Original Assignee: XY, LLC (Inguran LLC)
• Inventors: Seidel, George E., Herickhoff, Lisa A., Schenk, John L.
• Primary Examiner(s): Kim, Taeyoon
• Assistant Examiner(s): GOUGH, TIFFANY MAUREEN
• Application Number:
  US13/764,390
• Publication Number:
  US 20130149737A1
• Time in Patent Office: 1,289 Days
• US Class Current: 1/1
• CPC Class Codes: A01K 67/027 New or modified breeds of vertebrates
• CPC Class Codes: A01N 1/02 Preservation of living parts
• CPC Class Codes: A01N 1/0226 Physiologically active agents, i.e. substances affecting physiological processes of cells and tissue to be preserved, e.g. anti-oxidants or nutrients
• CPC Class Codes: A61D 19/02 for artificial insemination
• CPC Class Codes: A61D 19/04 for embryo transplantation
• CPC Class Codes: C12N 15/873 Techniques for producing new embryos, e.g. nuclear transfer, manipulation of totipotent cells or production of chimeric embryos
• CPC Class Codes: C12N 5/0612 sorting of gametes, e.g. according to sex or motility
• CPC Class Codes: C12Q 1/04 Determining presence or kind of microorganism; Use of selective media for testing antibiotics or bacteriocides; Compositions containing a chemical indicator therefor C12Q1/6897 takes precedence
• CPC Class Codes: G01N 1/30 Staining; Impregnating ; Fixation; Dehydration; Multistep processes for preparing samples of tissue, cell or nucleic acid material and the like for analysis
• CPC Class Codes: G01N 15/149 specially adapted for sorting particles, e.g. by their size or optical properties
• CPC Class Codes: G01N 33/52 Use of compounds or compositions for colorimetric, spectrophotometric or fluorometric investigation, e.g. use of reagent paper and including single- and multilayer analytical elements immunological elements G01N33/54386; involving labelled immunochemicals G01N33/58; for haemoglobin or occult blood G01N33/72
• CPC Class Codes: Y10T 436/101666 Particle count or volume standard or control [e.g., platelet count standards, etc.]
• CPC Class Codes: Y10T 436/108331 Preservative, buffer, anticoagulant or diluent

Abstract

Improved flow cytometer system particularly adapted to use for sex-selected sperm sorting include enhanced sheath fluid and other strategies which minimize stress on the sperm cells, including a 2.9 percent sodium citrate sheath solution for bovine species and a hepes bovine gamete media for equine species. Improved collection systems and techniques for the process are described so that commercial applications of sperms samples as well as the resulting animals may be achieved.

Claims

1. A method of isolating desired cells comprising the steps of:

a) supplying sperm cells to a nozzle of a flow cytometer;

b) supplying a sheath fluid comprising a citrate to the nozzle;

wherein the citrate is added to the sheath fluid prior to supplying the sheath fluid to the nozzle;

c) producing a stream containing the sperm cells from the nozzle;

d) sensing a property of sperm cells in the stream, the sensed property being indicative of a sex characteristic;

e) identifying X-chromosome bearing and/or Y-chromosome bearing sperm cells in the stream; and

f) collecting said X-chromosome bearing and/or Y-chromosome bearing sperm cells.

2. The method of claim 1, wherein the citrate comprises a chemical within the citric acid cycle.

3. The method of claim 1, wherein the sperm cells are stained with Hoechst 33342.

4. The method of claim 1, wherein the step of collecting sperm cells having the desired sex characteristic comprises charging drops based on the sensed properties of the sperm cells contained within those drops and deflecting those drops to an appropriate collector.

5. The method of claim 4, wherein the collector comprises a stream matched container.

6. The method of claim 1, wherein the sperm cells are present in a pre-sort environment prior to being supplied to the nozzle and wherein the sperm cells are present in a post-sort environment in a collector.

7. The method of claim 6, wherein the pre-sort environment and/or the post sort environment comprise a citrate.