Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving antigen-antibody binding – specific binding protein...
Patent
1980-12-10
1983-08-16
Padgett, Benjamin R.
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving antigen-antibody binding, specific binding protein...
424 85, 424 88, 435 4, 435 21, 435 25, 435 28, 435188, 436 71, 436518, 436524, 436808, 422 61, G01N 3392, G01N 3354
Patent
active
043992170
DESCRIPTION:
BRIEF SUMMARY
The invention relates to a process for the determination of serum lipoproteins as well as devices for implementing this process. More precisely, the process of the invention is intended for the determination of the protein components of serum lipoproteins.
Recent epidemiological studies on the risks of appearance of cardiovascular illnesses have shown the advantage of determining not only the global amount of serum lipoproteins but also and especially by making a distinction according to the group to which these lipoproteins belong and, within these groups, according to the type of apoprotein (C.sub.1, C.sub.2, C.sub.3, E, B, A.sub.1, A.sub.2). Traditionally, the three principal groups considered are the VLDL, LDL and HDL, these abbreviations corresponding to the English terms "very light density lipoprotein", "light density lipoprotein" and "high density lipoprotein". This designation refers to the way in which the separation is accomplished in ultracentrifugation separation techniques.
The importance of this distinction resides in the fact that close correlations have been able to be established, on the one hand, between the proportions of these types of lipoproteins and, on the other hand, the risk of appearance of the illnesses in question. Very succinctly, the risk is all the greater the higher the proportions of VLDL and LDL (and of apoprotein B) and, on the contrary, all the lower the higher the proportion of HDL.
For measuring the proportions of the different types of lipoproteins, several techniques have been previously proposed, which comprise separation of the different fractions of lipoproteins from the sample analyzed then the determination of the fractions thus separated. The principal difficulty with this kind of analysis resides in the separation of the different fractions. Thus, ultracentrifugation separation methods are not practicable for analyses carried out in a large number. For these routine analyses the only methods used at present comprise separation by selective precipitation. The accuracy of these methods rests particularly on the specificity of the precipitation carried out in earlier researches, and every effort has been made to perfect means improving this specificity. Appreciable improvements have been able to be obtained in this field by an appropriate choice of conditions and agents used for this precipitation. Furthermore, the traditional methods do not allow separation of each type of apoprotein within the same group. However, an additional increase of accuracy and sensitivity of measurement, while maintaining great facility of use and the possibility of determining the proportion of each apoprotein, was desirable.
The pesent invention proposed then providing means for effecting a determination of the different lipoproteins or fractions of lipoproteins, which is at one and the same time simple, very sensitive and accurate.
This aim was reached, in accordance with the invention, by using, for the determination of serum lipoproteins, a process combining immunological and enzymatic techniques and presenting the specificity of the one kind and the great sensitivity of the other.
According to the invention, the process consists in reacting, in conditions appropriate for the formation of a complex of the antibody-antigen type, on the one hand, a given amount of a specific antibody of the apoprotein(s) whose quantity it is desired to determine, this antibody being fixed on a solid support inert with respect to the reactive medium in which the antibody-antigen complex is formed and with respect to the medium in which the enzymatic activity is subsequently measured, on the other hand, the serum sample to be analyzed and a given amount of a compound for coupling the lipoprotein of the type of the one whose content is to be determined, or of several lipoproteins among which the one whose quantity is to be determined, with an enzyme which does not impair the formation of the antigen-antibody complex; with this latter accomplished, the reagents not fixed on the solid support phase are
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Carlson Lars A.
Holmquist Leif T.
Laboratoires Goella
Morkowitz M.
Padgett Benjamin R.
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