Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving virus or bacteriophage
Patent
1995-10-27
1999-01-26
Knode, Marian C.
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving virus or bacteriophage
435 71, 435 72, 435973, 436501, C12Q 170, G01N 3353, G01N 33566, G01N 33569
Patent
active
058637206
DESCRIPTION:
BRIEF SUMMARY
This application is the National Stage of International Application No. PCT/GB94/00298, filed Feb. 15, 1994, under 35 U.S.C. 371.
The present invention relates to a one-pot double serological assay for assessing the status of individuals potentially infected with diseases such as hepatitis B.
Hepatitis B is caused by the hepatitis B virus (HBV) and may be transmitted by blood transfusion (among other means). Screening of donated blood to detect every donation capable of transmitting HBV is highly desirable but presently very complex because a number of different tests need to be completed in order fully to characterise the infection status of the donor and thus to exclude those donations which may transmit the disease. It is now routine to screen donations for the surface antigen of HBV (designated HBsAg) and to discard all those giving a positive result (i.e. those from individuals with an established infection). This is, however, not sufficient to remove all potentially infective donations.
The donors whose samples pass the preliminary screening for HBsAg be divided into a number of categories as follows: in category 1 are distinguished by the absence of all HBV markers. Donors in category 2 have no serologically detectable markers but may be identified by the presence of genomic material from HBV using the polymerase chain reaction and hybridisation techniques. Donors in categories 3, 4 and 5 all show some serological markers of HBV infection but currently can be distinguished only on the basis of a complex sequence of tests.
In more detail, individuals in category 3 have cleared the HBsAg itself and mounted an immune response to the core antigen (HBcAg) but have not produced a protective antibody response to the surface antigen. They are therefore serologically identifiable by the presence of antibody against HBcAg (i.e anti-HBc) but low or absent anti-HBs. Individuals in category 4 have had a chronic infection and at some time would have been regarded as "carriers" of HBsAg; they lack anti-HBs but have high levels of anti-HBc. Individuals in category 5, having cleared the infection completely, have both anti-HBc and anti-HBs, some (hereafter category 5a) having very high levels of anti-HBs. Individuals in category 5a represent an important source of material for the production of hyperimmune globulin ("HBIg") which is valuable for passive immunisation against HBV infections. Identification of this subset of immune individuals is therefore important but at present requires further testing.
If these categories were to be distinguished on the basis of conventional single assays, a protocol such as the following would be required:
______________________________________ a) Test for HBsAg;
this is mandatory in many
blood transfusion programmes,
positive donations are
discarded.
b) Test for anti-HBc;
all positive donations are
quarantined.
c) Test positives
all positives
from test (b) with serum anti-HBs of over
for anti-HBs in a
0.1 I.U./ml are reinstated as
sensitive and donors (category 5), all others
quantitative are discarded (i.e. categories
test; 3 and 4).
d) Test positives
all those found with anti-HBs
from test (c) over 5 I.U./ml are candidates
for anti-HBs in an
for use inthe production of
insensitive mode;
HBIg (i.e. category 5a).
______________________________________
One objective of the present invention is to provide a "one-pot" assay which will enable donations from individuals in categories 3 and 4 to be excluded from use in blood transfusion and, preferably, also permits identification of individuals in category 5a with high levels of anti-HBs as opposed to those with moderate but safe levels of anti-HBs (hereafter category 5b).
In developing an assay to meet these objectives the present inventors have elucidated certain principles which may be applied to assays of other diseases where certain patterns of antibody or other markers might indicate infectivity or a serious illness.
The present invention therefore provides a one-pot double assay process which process c
REFERENCES:
patent: 5158869 (1992-10-01), Pouletty et al.
Journal of Medical Virology, vol. 6, 1980, New York, NY USA, pp. 323-332. R.S. Tedder et al. 'Contrasting patterns and frequency of antibodies to the surface, core and e antigens of hepatitis B virus in blood donors and in homosexual patients' see the whole document.
Journal of Virological Methods, vol. 11, 1985, Amsterdam NL, pp. 231-239. R.B. Ferns et al. 'Detection of both hepatitis B e antigen and antibody in a single assay using monoclonal reagnets'see the whole document.
Ratnam, S. and A.M. Tobin, "Comparative evaluation of commercial enzyme immunoassay kits for detection of hepatitis B seromarkers", J. Clin. Microbiol., 25:432-434, Feb. 1987.
Ekins and Chu, "Multianalyte microspot immunoassay--microanalytical `compact disk`of the future", Clin. Chem. 37/11, 1955-1967, 1991.
Brunback Brenda G.
Cooper Iver P.
Knode Marian C.
University College London
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