Immobilization of enzymes on human tissue or erythrocytes

DETAILED DESCRIPTION I claim: 1.
A method for immobilizing an enzyme on tissue or erythrocytes comprising the steps of reacting the enzyme having nucleophilic groups selected from the group consisting of amino, carboxyl and tyrosine groups with a compound containing an electrophilic group selected from the group consisting of .
alpha.
, .
beta.
unsaturated carbonyls, alkyl halides, and thiol to form an enzyme derivative, and then immobilizing the enzyme on the tissue or erythrocyte having nucleophilic groups selected from the group consisting of sulfhydryl, amino, imidazole, carboxylato and tyrosine groups by a coupling reaction between electrophilic groups of the derivative and nucleophilic groups in the tissue or erythrocyte.
2.
The method as claimed in claim 1 in which the nucleophilic group with which the enzyme derivative is interbonded in the tissue or erythrocyte is a sulfhydryl group.
3.
The method as claimed in claim 1 in which the coupling compound in one containing free functional groups selected from the group consisting of free carboxyl groups, free amino groups, free aromatic amino groups, isocyanate groups, imidoester groups, acylating and alkylating groups.
4.
The method as claimed in claim 1 which includes the step of reacting the enzyme or compound with an activator selected from the group consisting of chloro formate, carbodiimides and isoxazolium salt, or nitrous acid for aromatic amino groups prior to reaction of the compound with the enzyme to form the derivative.
5.
The method as claimed in claim 4 in which the isoxazolium salts react with the enzyme to form an enol ester derivative of the enzyme.
6.
The method as claimed in claim 4 in which the activator is isoxazolium salt.
7.
The method as claimed in claim 1 in which the enzyme derivative is formed by reaction of isobutylchloro formate as an activator.
8.
The method as claimed in claim 4 in which the activator is Woodward's reagent K.




Description:
This invention relates to the immobilization of enzymes and it relates more particularly to the immobilization of enzymes on human erythrocytes to retard their elimination and/or to enhance their biological activity