Bio-tech Imaging, Inc.

The Avriel Group, Amcas Division Inc.

Robert A Hallowitz

Chester F King

Susan M Young

The present invention relates to cells which have improved receptivity to viruses which are capable of infecting them. Receptivity to such viruses is improved by selecting cells from a population which express the receptor(s) that enable a virus to attach to the cell and gain entry into it. Any combination of viruses and host cell lines can be used. In a preferred embodiment, the present invention relates to improving receptivity or infectivity of a cell line which can be infected with an immunodeficiency virus, such as HIV-1. Especially preferred embodiments of the invention relate to methods of improving (or assaying for) the infectivity for HIV-1 in a HIV-1 receptive cell line, preferably a continuous cell line transformed with DNAs coding for expressible CD4 and expressible HIV-1 coreceptor, comprising, in any effective order, a) isolating the cells expressing CD4 and an HIV-1 coreceptor on their cell surface; b) contacting the isolated cells with HIV-1 under conditions effective for the HIV-1 to infect the cells; and c) detecting the number of cells infected with HIV-1, thereby assaying for infectivity of HIV-1. This method facilitates the measurement of true infectivity and infectivity reduction values by quantifying the percentage of infected cells in the population of specific cells capable of being infected by virus, rather than in a population of mixed cells, only some which are capable of being infected.

Methods of improving infectivity of cells for viruses

A microorganism detecting apparatus comprising a fluorescence microscope section which is furnished with a motor-driven xyz stage assembly for placing thereon a slide sample subject to fluorescent staining. An illumination subsystem for projecting excitation light of predetermined wavelength on the sample and a filter assembly for emission, which limits the band of frequencies emitted to a video camera which captures the images of fluorescent stained microorganisms from the sample slide mounted on the xyz stage assembly. A computer reads the output value of the band-limited signal from the video camera and processes the read output. A subsystem device has controllers which drive the xyz stage assembly so as to permit the microorganism sample to be scanned over its whole area. Each image is displayed on a video display monitor and detected microbes in the sample are stored on a hard drive and in an imaging archive system to permit verification or reexamination.

Method of and apparatus for automating detection of microorganisms

Fluorometric immunological assay method for detection of HIV-1 infection in which Murine anti-gp120 monoclonal antibodies coupled to paramagnetic microspheres (14) and Fluorescein conjugated anti-gp120 polyclonal antibodies IgG (16) are incubated in a few drops of whole blood diluted in 0.5 cc phosphate buffered saline (10). After incubation for 5 minutes, the HIV-infected peripheral blood lymphocytes (18) will be coated with both the Murine anti-gp120 monoclonal antibodies coupled to paramagnetic microspheres (14) and Fluorescein conjugated anti-gp120 polyclonal antibodies IgG (16) at exposed gp120 antigens (20) binding sites. At the time of measurement said HIV- infected peripheral blood lymphocytes (18) will be pulled against the wall of the measurement vessel by means of a magnetic gradient (26). The cells adhering to the vessel wall are illuminated at 488 nm monochromatic light by a focused light source (28) and the resultant emitted fluorescence is imaged, measured and recorded.

Reagent system for detecting HIV-infected peripheral blood lymphocytes

The invention is a device and a method for collecting samples mixing the samples with test reagents, and acting as a container in which the mixture can be incubated and the test reaction viewed by microscope or imaging device. This device enables an entire test to be performed in one simple step without complicated handling procedures. The device consists of a cartridge with a well with micro-lances imbedded in the bottom of the well and an overlying micro-baggy containing a mixture of reagents. There are two reagents present in the micro-baggy: the first consisting of antibodies coupled to paramagnetic microspheres and the second consisting of antibodies coupled with a fluorochrome. A test subject presses down onto the micro-baggy and at the same time punctures his/her finger or thumb on the micro-lances. Once the finger has been lanced, breaking the micro-baggy, the reagents mix with the test subject's blood. The well is then covered by a clear mylar strip. After incubation the cartridge is moved to a viewing platform. A magnetic gradient is applied to the side of the well causing the magnetic separation of antibodies bound to infected peripheral blood lymphocytes from uninfected peripheral blood lymphocytes. A focused light source is applied to the well, causing antibodies bound to infected peripheral blood lymphocytes if any are present, to glow at the specific emmision frequency determined by the specific fluorochome. The test subject can be identified by a bar code strip attached to the cartridge.

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Gaithersburg, MD, United States of America

Robert A Hallowitz