Reverse immunoaffinity chromatography purification method

Claim 1

A method for the purification of a protein component of a biological fluid, said method comprising: (a) raising antibodies to all of the antigenic substances other than said protein, commonly present, as impurities, in a crude preparation of said protein, and purifying said antibodies; (b) preparing an immunoadsorbent complex by linking said antobodies against said impurities to a solid adsorbent suitable for use in column chromatography; (c) processing a preparation of said protein containing said impurities through a chromatography column containing said immuoadsorbent, thereby causing selective adsorption of said impurities in one step and exclusion of said protein in the effluent; and (d) recovering said purified protein from said effluent.

Claim 2

A method for the purification of a protein component of a biological fluid, said method comprising: (a) raising antibodies to all of the antigenic substances other than said protein, commonly present, as impurities, in a crude preparation of said protein, and purifying said antibodies; (b) preparing an immunoadsorbent complex by linking said antobodies against said impurities to a solid adsorbent suitable for use in column chromatography; (c) processing a preparation of said protein containing said impurities through a chromatography column containing said immuoadsorbent, thereby causing selective adsorption of said impurities in one step and exclusion of said protein in the effluent; and (d) recovering said purified protein from said effluent.

Claim 11

A method for the purification of a ptorein component of a biological fluid said method comprising: (a) concentration said biological fluid; (b) centrifuging said concentrate to remove insoluble impurities present in said fluid; (c) processing said centrifuged concentrate through a crosslinked neutral gel chromatographic column, said gel containing a hydrophobic group, whereby non-hydrophobic contaminants present in said centrifuged concentrate are exluded and hydrophobic constituents of said centrifuged concentrate are subsequently recovered by elution from said column; (d) raising antibodies to all of the antigenic substances other than said protein commonly present, as impurities, in crude preparations of said protein by injecting in laboratory animals a quantity of a curde preparation of said protein sufficient to induce an immune response in said animals, obtaining specific anitsera from said animals and separating said antigenic impurity antibodies from said specific antisera; (e) linking immunoglobulins of said specific antisera, after separation of said antigenic impurity antibodies therefrom, to a first solid adsorbent suitable for use in column chromatography; (f) processing said constituents of said concentrate recovered by elution in step (c) through said first immunoadsorbent, whereby the bulk of the impurites remaining in said step (c) concentrate are excluded in the effluent and recovering, as a product, the remaining consitutents of said step (c) concentrate by elution; (g) linking said antigenic impurity antibodies to a second solid adsorbent support suitable for use in column chromatography; (h) processing said step (f) product through said second column, wherein the remaining impurites of said protein adsorb to their antibodies and the protein is exclused in the effluent; and (i) recovering said purified protein from said second column effluent.

Claim 12

A method for the purification of a ptorein component of a biological fluid said method comprising: (a) concentration said biological fluid; (b) centrifuging said concentrate to remove insoluble impurities present in said fluid; (c) processing said centrifuged concentrate through a crosslinked neutral gel chromatographic column, said gel containing a hydrophobic group, whereby non-hydrophobic contaminants present in said centrifuged concentrate are exluded and hydrophobic constituents of said centrifuged concentrate are subsequently recovered by elution from said column; (d) raising antibodies to all of the antigenic substances other than said protein commonly present, as impurities, in crude preparations of said protein by injecting in laboratory animals a quantity of a curde preparation of said protein sufficient to induce an immune response in said animals, obtaining specific anitsera from said animals and separating said antigenic impurity antibodies from said specific antisera; (e) linking immunoglobulins of said specific antisera, after separation of said antigenic impurity antibodies therefrom, to a first solid adsorbent suitable for use in column chromatography; (f) processing said constituents of said concentrate recovered by elution in step (c) through said first immunoadsorbent, whereby the bulk of the impurites remaining in said step (c) concentrate are excluded in the effluent and recovering, as a product, the remaining consitutents of said step (c) concentrate by elution; (g) linking said antigenic impurity antibodies to a second solid adsorbent support suitable for use in column chromatography; (h) processing said step (f) product through said second column, wherein the remaining impurites of said protein adsorb to their antibodies and the protein is exclused in the effluent; and (i) recovering said purified protein from said second column effluent.