PROCEDE DE PURIFICATION DE LA .beta.1-GLYCOPROTEINE SPECIFIQUE AUX FEMMES ENCEINTES

PROCESS FOR THE PURIFICATION OF THE PREGNANCY-SPECIFIC .beta.1-GLYCOPROTEIN

Abrégé anglais

PROCESS FOR THE PURIFICATION OF THE PREGNANCY-SPECIFIC .beta.1-GLYCO-
PROTEIN
Abstract of the disclosure:
Process for the purification of the pregnancy-specific .beta.1-glyco-
protein, which comprises contacting an aqueous phosphate-con-
taining solution which contains the pregnancy-specific .beta.1-glyco-
protein with hydroxyl-apatite, separating the hydroxyl-apatite
from the solution and isolating the SP1 from the solution.

Revendications

THE EMBODIMENTS OF THE INVENTION IN WHICH AN EXCLUSIVE
PROPERTY OR PRIVILEGE IS CLAIMED ARE DEFINED AS FOLLOWS:
1. A process for the purification of the pregnancy-
specific .beta.1-glycoprotein in which an aqueous phosphate-contain-
ing solution containing the pregnancy-specific .beta.1-glycoprotein
is contacted with hydroxyl-apatite, the hydroxyl-apatite is
separated from the solution and the pregnancy-specific .beta.1-
glycoprotein is isolated from the solution.
2. A process as claimed in claim 1 in which the pro-
cess is carried out in a pH range of from 5 to 8.5 and a molar-
ity of the phosphate of from 0.001 to 0.01.
3. A process as claimed in claim 1 in which the pro-
cess is carried out in a sodium or potassium phosphate solu-
tion in the pH range of from 6 to 8 and a molarity of the phos-
phate of 0.005.

Description

~065307
German Patent Application laid open to public inspection
DOS 21 57 610 describes a glycoprotein which occurs in the pla-
centa and is called "Pregnancy-specific ~l-glycoprotein" (herein-
after referred to as SPl). Said Patent Application furthermore
describes a process for the isolation of this glycoprotein.
The present invention relates to an adsorption process which
permits further purification of the pregnancy-specific Bl-glyco-
protein obtained by the above-mentioned process with a purity of
60 to 95% and to obtain it in practically pure form.
The process of the invention comprises contacting an aqueous,
phosphate-containing solution which contains the pregnancy-spe-
cific ~l-glycoprotein with hydroxy-apatite, separating the
hydroxyl-apatite from the solution and isolating the SPl from
the solution. The adsorption conditions are so chosen that the
pregnancy-specific ~l-glycoprotein is not adsorbed but that any
impurities still present are bound to the hydroxyl-apatite.
The adsorption is carried out in a phosphate-containing aqueous
medium, advantageously in a phosphate-containing buffer such as
potassium phosphate buffer or sodium phosphate buffer, in a pH-
range of from 5 to 8.5, preferably 6 to 8, and a molarity of the
phosphates of from 0.001 to 0.01, preferably 0.005.
The adsorption of the impurities may be carried out according
to a batch process or as column chromatography.
If the above-mentioned conditions are maintained, the
pregnancy-specific Bl-glycoprotein is obtained by this process
in a purity of ~99%.
The purity of the preparation is advantageously tested by
immunological processes using antisera which contain antibodies
- 2 -
q~

~065307
against the pregnancy-specific ~l-glycoprotein and some or
several antigens which occur in the extract of placentas, in
the blood or urine of pregnant women.
The following Example illustrates the invention.
EXAMPLE:
For highly purifying pregnancy-specific gl-glycoprotein,
50 mg of the preparation having a purity of 60 to 95% were
dissolved 10 ml of 0.005 m sodium phosphate buffer (pH 6.0)
and introduced in a column (2 x 8 cm) filled with hydroxyl-
apatite (Messrs. Serva, Heidelberg) and which had previously
been equilibrated with a 0.005 m phosphate buffer (pH 6.8).
During elution of the column with 0.005 m phosphate buffer, the
pregnancy-specific Bl-glycoprotein migrated through the
column, whereas the impurities were adsorbed on the hydroxyl-
apatite. The protein-containing eluate contained the pregnancy-
specific Bl-glycoprotein in pure form; in order to remove the
salts, it was dialyzed against 10-times the volume of water
and subsequently lyophilized. Yield: about 40 m of pregnancy-
specific ~l-glycoprotein. The purity was found to be > 99%,
i.e. by immunological methods none of the impurities found
before the purification operation could be detected.
~,,