Device for monitoring cells

• US 6,395,506 B1
• Filed: 06/29/1999
• Issued: 05/28/2002
• Est. Priority Date: 04/18/1991
• Status: Expired due to Term

• Current Assignee: Becton, Dickinson & Co
• Original Assignee: Becton, Dickinson & Co
• Inventors: Beaty, Patrick Shawn, Pitner, J. Bruce, Burrell, Gregory J., Guarino, Richard D., Foley, Timothy G. Jr., Stitt, David T., Hemperly, John Jacob, Wodnicka, Magdalena
• Primary Examiner(s): GITOMER, RALPH J
• Application Number:
  US09/342,720
• Time in Patent Office: 1,064 Days
• Field of Search: 435/29,34,69.1,32,817,968 436/172,800
• US Class Current: 435/32
• 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: C12Q 1/18 Testing for antimicrobial activity of a material
• CPC Class Codes: C12Q 2304/44 Oxygen
• CPC Class Codes: Y02A 90/10 Information and communication technologies [ICT] supporting adaptation to climate change, e.g. for weather forecasting or climate simulation

Abstract

The present invention relates to methods for detection and evaluation of metabolic activity of eukaryotic and/or prokaryotic cells based upon their ability to consume dissolved oxygen. The methods utilize a luminescence detection system which makes use of the sensitivity of the luminescent emission of certain compounds to the presence of oxygen, which quenches (diminishes) the compound'"'"'s luminescent emission in a concentration dependent manner. Respiring eukaryotic and/or prokaryotic cells will affect the oxygen concentration of a liquid medium in which they are immersed. Thus, this invention provides a convenient system to gather information on the presence, identification, quantification and cytotoxic activity of eukaryotic and/or prokaryotic cells by determining their effect on the oxygen concentration of the media in which they are present.

Claims

1. A method for determining the effects of at least one drug, toxin or chemical on respiring mammalian or insect cells comprising:

(i) preparing a liquid media broth of said mammalian or insect cells;

(ii) contacting said broth with a sensor composition which comprises a luminescent compound that exhibits a change in luminescent property, when irradiated with light containing wavelengths which cause said compound to luminesce, upon exposure to oxygen, wherein the presence of the sensor composition is non-destructive to the mammalian or insect cells;

(iii) admixing with said broth a quantity of said drug, toxin or chemical;

(iv) irradiating said sensor composition with light containing wavelengths which cause said luminescent compound to luminesce;

(v) measuring or visually observing the change in luminescent property from said luminescent compound while irradiating said sensor compound with said light; and

(vi) comparing said measurement to that of a control wherein said control is selected from the group consisting of;

a reagent control not in contact with respiring mammalian or insect cells or the drug, toxin or chemical;

a reagent control in contact with respiring mammalian or insect cells but not in contact with the drug, toxin or chemical and a calculated threshold, wherein a change in luminescent property relative to the control is indicative of cytotoxicity of the drug, toxin or chemical to the mammalian or insect cells; and

(vii) in the even that no such change is measured or observed, repeat steps (iv), (v) and (vi), as needed, to determine the effects of the drug, toxin or chemical on the respiring mammalian or insect cells.

2. The method of claim 1 wherein said luminescent compound is contained within a matrix which is relatively impermeable to water and non-gaseous solutes, but which has a high permeability to oxygen.

3. The method of claim 2 wherein said matrix is a rubber or plastic matrix.

4. The method of claim 2 wherein said matrix is a silicone rubber matrix.

5. The method of claim 2 wherein said luminescent compound is adsorbed on solid silica particles.

6. The method of claim 1 wherein said luminescent compound is a tris-4,7-diphenyl-1,10-phenanthroline ruthenium (II) salt.

7. The method of claim 6 wherein said luminescent compound is tris-4,7-diphenyl-1,10-phenanthroline ruthenium (II) chloride.

8. The method of claim 1 wherein said luminescent compound is a tris-2,2′

-bipyridyl ruthenium (II) salt.

9. The method of claim 8 wherein said luminescent compound is tris-2,2′

-bipyridyl ruthenium (II) chloride hexahydrate.

10. The method of claim 1 wherein said luminescent compound is 9,10-diphenyl anthracene.

11. The method of claim 1 wherein said broth is isolated from atmospheric oxygen.

12. The method of claim 1 wherein said broth is exposed to atmospheric oxygen.

13. The method of claim 1 wherein, in step (ii), the liquid media is also contacted with an effective concentration of one or more biomaterials, selected from the group consisting of laminin, collagen IV, entactin, heparan sulfate proteoglycans, defined mammalian and insect cell growth factors, and matrix metalloproteinases, said concentration being effective for promoting or enabling cellular growth and respiration.

14. The method of claim 13 wherein said biomaterial is Matrigel.

15. The method of claim 1 wherein, in step (ii), the liquid media is also contacted with an effective concentration of one or more extracellular matrices, said concentration being effective for promoting or enabling cellular growth and respiration.

16. The method of claim 15 wherein said extracellular matrix is collagen.

17. The method of claim 1 wherein, in step (ii), the liquid media is contacted with an effective concentration or one or more additives or coating substances, selected from the group consisting of penicillin, streptomycin, fungizone, non-essential amino acids, sodium pyruvate, and fetal bovine serum, said concentration being effective for promoting or enabling cellular growth and respiration.