A new analytical method exploiting molecular competition between two solid phases has been ideated for the simultaneous, quantitative detection of different cellular and molecular markers. The method uses 2 new shaped immunosorbent solid phases: the 1st device is the microstrip bearing 3 maxi-wells – each corresponding to 4 wells in a standard platform - in which are immerged 4 ogival pins protruding from a rod, that constitute the 2nd device. The method has been designed to set up a ready-to-use diagnostic kit for the quick diagnosis of tuberculosis in humans and of tuberculosis and early-stage paratuberculosis diagnosis in bovines, through detection of surface cell markers of infection-sensitized helper and cytotoxic T lymphocytes. Heparinized total blood samples are analyzed as follows: after the adsorbtion of T lymphocytes to the maxi-well surface, a competitive reaction starts in the presence of sensitized T lymphocytes when biotin-conjugate TB antigens are added in the maxi-well and the ogival pins, each coated with a specific monoclonal antibody directed to one of the added TB antigens, are immersed in the maxi-well. The OD observed after the incubation of each pin in a well of a standard platform containing avidin-HRP conjugate, followed by incubation in TMB, is inversely proportional to the amount of the respective T-cells surface receptors. The kit doesn’t require laboratory facilities to carry out samples analyses and can be easily robotized to increase the process performances.
Setting Up a New Platform for Fast and Portable Diagnosis of Tuberculosis
MUSCI, Giovanni;MAZZEO, Alessandra
2011-01-01
Abstract
A new analytical method exploiting molecular competition between two solid phases has been ideated for the simultaneous, quantitative detection of different cellular and molecular markers. The method uses 2 new shaped immunosorbent solid phases: the 1st device is the microstrip bearing 3 maxi-wells – each corresponding to 4 wells in a standard platform - in which are immerged 4 ogival pins protruding from a rod, that constitute the 2nd device. The method has been designed to set up a ready-to-use diagnostic kit for the quick diagnosis of tuberculosis in humans and of tuberculosis and early-stage paratuberculosis diagnosis in bovines, through detection of surface cell markers of infection-sensitized helper and cytotoxic T lymphocytes. Heparinized total blood samples are analyzed as follows: after the adsorbtion of T lymphocytes to the maxi-well surface, a competitive reaction starts in the presence of sensitized T lymphocytes when biotin-conjugate TB antigens are added in the maxi-well and the ogival pins, each coated with a specific monoclonal antibody directed to one of the added TB antigens, are immersed in the maxi-well. The OD observed after the incubation of each pin in a well of a standard platform containing avidin-HRP conjugate, followed by incubation in TMB, is inversely proportional to the amount of the respective T-cells surface receptors. The kit doesn’t require laboratory facilities to carry out samples analyses and can be easily robotized to increase the process performances.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
