Apparatus for the in situ hybridization of slide-mounted cell sa

Details Apparatus for the in situ hybridization of slide-mounted cell sa Apparatus for the in situ hybridization of slide-mounted cell sa

1992-01-14

1993-07-27

Housel, James C.

Chemistry: molecular biology and microbiology

Apparatus

Bioreactor

422 67, 435316, 435809, 436 46, 436 47, 436 49, 436 50, 436174, C12Q 168, G01N 130

Patent

active

052310299

DESCRIPTION:

BRIEF SUMMARY
This invention relates to in situ hybridization and in particular to apparatus for effecting hybridization of cell samples carried on slides.
A powerful technique which has been developed for detecting particular sequences of nucleic amino-acids in a segment of DNA is known as in situ hybridization. This technique uses a chemically or radioactively labelled "probe" to hybridize the DNA of tissue samples in situ, the samples which become labelled can then be detected by the physical or chemical properties of their chemical label or their radioactivity. Because of the risk to the health of the operator through the use of radioactive materials, chemically labelled probes may be preferred.
The process of in situ hybridization requires the treatment of samples on a microscope slide, for example, with measured small quantities, say 50 .mu.liters, of reagents such as enzymes. The reagents are applied by pipette or other hand-held dispenser to the slide and then spread evenly over the surface. These pre-hybridization reagents are then washed off in, say, phosphate buffered solution (PBS) and a hybridization "cocktail" of further reagents applied in the same way as the first reagents. The samples are then covered by a glass cover-slip or a film of inert plastics material to prevent them drying out during incubation which may extend for a period between say 30 minutes and 16 hours or even more. After that the cover-slip or film may be removed, the slide washed in, say, phosphate buffered solution (PBS), further reagents applied and incubation carried out for a second period. There are many different "recipes" for in situ hybridization, some involving several treatments with reagents and several incubations. It is evident that the process must be performed carefully by skilled personnel and that it is very time-consuming. It is therefore desirable to reduce the labour content of the process without compromising the care and accuracy of the treatment of the samples.
It is an object of the present invention to provide a machine which is capable of performing in situ hybridization on slide-mounted tissue samples.
According to the present invention there is provided apparatus for the in situ hybridization of slide-mounted cells samples in which slides bearing samples are brought successively to a station to be sprayed with one or more fluid reagents by a spray means and thereafter are subjected to incubation to further the process,
the spray means including a reservoir into which a fluid reagent can be fed and a spray head to which the fluid reagent can be passed from the reservoir for spraying on a sample on a slide,
wherein the apparatus includes at least one container for containing a fluid reagent, pipe means leading from the or each container to the reservoir, valve means selectively effective to block and to permit flow along the pipe means, and means for selectively establishing an air pressure difference between the or each container and the reservoir, the arrangement being such that the air pressure difference can cause fluid to flow from a particular container into the reservoir when the corresponding valve means is such as to permit flow along the pipe means, the valve means being operated to permit a predetermined volume of a fluid reagent to be fed to the reservoir.
The feeding of the fluid into the reservoir and the passing of the fluid from the reservoir to the spray head may be effected at least partly by air pressure. The air pressure may be provided by a higher than atmospheric pressure applied to the fluid to cause it to flow to a region held at atmospheric pressure or it may use atmospheric pressure to cause the fluid to flow to a region where a pressure below atmospheric pressure is maintained.
The apparatus may include a container for containing fluid subject to atmospheric pressure, with the pipe means leading from the container to the reservoir, and the valve means selectively effective to block and to permit flow along the pipe means, and a suction pump selectively connectible to draw air from th

REFERENCES:
patent: 3650437 (1972-03-01), Binnings et al.
patent: 3875899 (1975-04-01), Clements
patent: 4004550 (1977-01-01), White et al.
patent: 4013038 (1977-03-01), Rogers et al.
patent: 4043292 (1977-08-01), Rogers et al.
patent: 4543236 (1985-09-01), Von Gise
patent: 5009185 (1991-04-01), Stokes et al.

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