Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving cholesterol
Reexamination Certificate
1998-07-14
2001-04-10
Gitomer, Ralph (Department: 1623)
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving cholesterol
C422S051000, C436S013000
Reexamination Certificate
active
06214570
ABSTRACT:
The invention concerns a method for the quantitative determination of HDL (
H
igh
D
ensity
L
ipoprotein) in biological fluids and an agent suitable therefor.
Total cholesterol in blood, plasma or serum is one of the best known parameters for assessing the extend of risk of a coronary heart disease. However, the concentration of total cholesterol is only of limited value for the assessment of individual risk. The measurement of the cholesterol in the lipoproteins of low density (
L
ow
D
ensity
L
ipoproteins=LDL) on the one hand and in the lipoproteins of high density (
H
igh
D
ensity
L
ipoproteins=HDL) on the other hand is more meaningful. Epidemiological and clinical studies have shown that there is a positive correlation between LDL cholesterol and coronary heart disease and a negative correlation between EDL cholesterol and coronary heart disease.
As a close approximation the determination of the HDL as well as the total cholesterol is sufficient for an assessment of risk. This course is preferably followed at present in diagnostic practice.
The other lipoprotein classes (LDL, VLDL, chylomicrons) which are present have to be separated in order to determine HDL cholesterol separately. The potential methods of separation are based on differences in the flotation densities (sequential flotation or equilibrium sedimentation, both in the ultracentrifuge), on different surface charges (electrophoreses on paper or agarose as carrier) or on differences in the apolipoproteins (immunochemical methods using specific antibodies). All these methods of separation are expensive, tine-consuming and not established in routine laboratories. Precipitation reactions (in Monographs on Atherosclerosis, Vol. 11 (1982), Clackson, T. B., Kritchevsky, D., Pollak, O. J. eds; Lipoprotein Precipitation, Burstein, M., Legmann, P. and in Meth. in Enzymology, Vol. 129 (1986)) whose specificity depends on the particle dimension and the surface charge are cheap, relatively easy to handle and therefore widespread. Polymeric substances serve as precipitation reagents, and these are usually of a polyanionic nature. Polymers which are uncharged are also suitable. The polyanions usually need bivalent cations in order to develop their precipitating effect, while the uncharged polymers do not require them.
The experimental procedures and the concentrations which are used for the combined precipitation agents are designed to quantitatively precipitate all lipoproteins except HDL, to separate these precipitates from the liquid fraction of the sample in a suitable manner and subsequently to quantify the HDL in the liquid fraction of the sample by means of a cholesterol assay. For this, depending on how the test is carried out, a defined volume of precipitating agent (in suitable concentrations) is mixed intensively with a defined volume of the sample to be determined. It is the state of the art to allow a reaction time of at least 10 minutes for the precipitation and only after this time interval has elapsed to separate sedimented non-HDL lipoprotein precipitates and HDL remaining in the liquid fraction by centrifugation. The centrifugation step also needs some time.
This method is much too time-consuming for a routine test. In addition, centrifugation steps with subsequent separation of the supernatant need complicated additional equipment and a transfer step. The required pipetting procedure in which a defined amount of supernatant is taken is, in addition, a source of error which can lead to less precise measurements.
It is therefore the object of the invention to avoid the disadvantages of the state of the art and to provide a method for the separation of non-HDL lipoproteins from biological fluids which can be carried out more rapidly and without complicated additional equipment and which allows a more rapid and a simpler HDL cholesterol determination.
The object is achieved by a method for the separation of non-HDL lipoproteins from biological fluids, in which the biological fluid containing non-HDL lipoproteins is applied onto a carrier through which liquids can flow and which contains a precipitating agent for non-HDL lipoproteins. The invention also provides an agent for the separation of non-HDL lipoproteins and a rapid diagnostic agent which contains this agent.
REFERENCES:
patent: 4604264 (1986-08-01), Rothe et al.
patent: 4746605 (1988-05-01), Kerscher et al.
patent: 5426030 (1995-06-01), Rittersdorf et al.
patent: 5580743 (1996-12-01), Rittersdorf et al.
patent: 5786164 (1998-07-01), Ritterdorf et al.
Buecker Klaus
Deneke Ulfert
Goebbert Uwe
Hiller Gerhard
Merdes Hartmut
Fulbright & Jaworski LLP
Gitomer Ralph
Roche Diagnostics GmbH
LandOfFree
Method for separating non-HDLs from HDLs, and determining,... does not yet have a rating. At this time, there are no reviews or comments for this patent.
If you have personal experience with Method for separating non-HDLs from HDLs, and determining,..., we encourage you to share that experience with our LandOfFree.com community. Your opinion is very important and Method for separating non-HDLs from HDLs, and determining,... will most certainly appreciate the feedback.
Profile ID: LFUS-PAI-O-2533020