Automated Staining and Decolorization of Biological Material

Abstract

An improved method and apparatus for staining samples of biological material for accurate analysis of the sample. Biological material is applied to a substrate, such as a microscope slide. The biological specimen is then stained with a selected staining composition, which may be gentian violet for a Gram's Stain analysis. The stained biological material is at least partially decolorized and the level of decolorization is analyzed optically. If necessary, the decolorizing step and the optical analysis steps are repeated until a selected level of decolorization is obtained.

1 . (canceled) 
   
   
       2 . Apparatus for automatically staining a biological specimen on a microscope slide, the apparatus comprising:
 a microscope slide carrier;   a plurality of fluid reservoirs, each containing a fluid, the fluids including at least a stain and a decolorizer;   a controller coupled to the fluid reservoirs to control flow of the fluids onto the biological specimen on the microscope slide;   a collector configured to at least temporarily collect decolorizer that runs off the biological specimen;   an electro-optical detector coupled to the controller and arranged to detect a level of decolorization in the decolorizer collected in the collector by comparing a characteristic of the collected decolorizer to a selected threshold that indicates an adequate level of decolorization in the biological specimen.   
   
   
       3 . The apparatus according to  claim 2 , wherein the stain comprises Gentian violet and iodine. 
   
   
       4 . The apparatus according to  claim 2 , wherein the decolorizer is acid alcohol. 
   
   
       5 . The apparatus according to  claim 2 , wherein the electro-optical detector further comprises:
 a source of radiation arranged to irradiate the collector and collected decolorizer with radiation of at least one selected wavelength that is absorbed by stain contained in the collected decolorizer;   a radiation detector configured to detect a degree of absorption of the radiation of selected wavelength by the collected decolorizer, the detector coupled to the controller for comparison of the degree of absorption with the selected threshold.   
   
   
       6 . The apparatus according to  claim 5 , wherein the selected wavelength of the radiation is that corresponding to a violet portion of the visible spectrum. 
   
   
       7 . The apparatus according to  claim 5 , wherein the radiation detector is a photodiode. 
   
   
       8 . Apparatus for automatically staining a biological specimen on a microscope slide, the apparatus comprising:
 a microscope slide carrier;   a plurality of fluid reservoirs, including at least a reservoir of stain fluid and a reservoir of decolorizer;   a controller coupled to the fluid reservoirs to control dispensation of at least the stain fluid and the decolorizer onto the biological specimen on the microscope slide on the slide carrier;   a collector configured to at least temporarily collect decolorizer that runs off the biological specimen;   an electro-optical detector coupled to the controller and arranged to detect a level of decolorization in the decolorizer collected in the collector by comparing a characteristic of the collected decolorizer to a selected threshold that indicates an adequate level of decolorization in the biological specimen.   
   
   
       9 . The apparatus according to  claim 8 , wherein the stain comprises Gentian violet and iodine, and each is contained in a separate fluid reservoir. 
   
   
       10 . The apparatus according to  claim 8 , wherein the decolorizer is acid alcohol. 
   
   
       11 . The apparatus according to  claim 8 , wherein the electro-optical detector further comprises:
 a source of radiation arranged to irradiate the collected decolorizer with radiation of at least one selected wavelength that is absorbed by stain contained in the collected decolorizer;   a radiation detector configured to detect a degree of absorption of the radiation of selected wavelength by the collected decolorizer, the detector coupled to the controller for comparison of the degree of absorption with the selected threshold.   
   
   
       12 . The apparatus according to  claim 11 , wherein the selected wavelength of the radiation is that corresponding to a violet portion of the visible spectrum. 
   
   
       13 . The apparatus according to  claim 11 , wherein the radiation detector is a photodiode.