# US 4,396,600 A · Adult schistosome worm-derived antigenic substance and method of obtaining same > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 4,396,600 A - Document ID: US4396600A - Status: Granted - Kind: A - Application number: 06/217,575 - Filed: 1980-12-18 - Granted: 1983-08-02 - Earliest claimed: 1980-12-18 - Assignees: Gus Gallucci; Mike Gallucci; Michael Gallucci, Jr.; Don Lower - Inventors: Luigi Messineo; Mauro Scarpin - Web page: https://patentgrep.com/patent/US4396600A ## CPC - A61K 39/0003: {Invertebrate antigens} - Y02A 50/30: Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change - Y10S 530/806: Antigenic peptides or proteins - Y10S 530/855: Proteins from animals other than mammals or birds - A61K: PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES (devices or methods specially adapted for bringing pharmaceutical products into particular physical or administering forms A61J3/00; chemical aspects of, or use of materials for deodorisation of air, for disinfection or sterilisation, or for bandages, dressings, absorbent pads or surgical articles A61L; soap compositions C11D) - Y02A: TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE - Y10S: TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS ## Abstract An extract of adult Schistosome mansoni worms, obtained by incubation in 0.15 M sodium chloride-sodium phosphate buffer (pH 6.8), contains protein, carbohydrates, and nucleic acid and/or by-products of the latter component and resolves into four major fractions by gel chromatography in G-100 and G-200 Sephadex columns. Immunodiffusion tests with rabbit anti-total extract serum reveal three precipitation lines corresponding to fractions I and II, and one with III or IV. Rabbits immunized with this total extract are found to be totally or partially (at least 77%) resistant to a challenge infection. The saline extract antigenic material is an effective vaccine for the treatment and immunization of schistosomiasis and other schistosome infections. ## Claims ### Claim 1 An antigen-containing extractant capable of developing an immune reaction in mammals to infection by schistosomes comprising a saline extract obtained from live adult schistosomes by the steps of: collecting fresh, living adult schistosomes from a host without destroying the schistosomes; rinsing the fresh, living adult schistosomes with saline solution to remove only blood and non-worm solids but without removing worm by-products; extracting antigenic material from the rinsed adult schistosomes in a saline extracting solution for a period of at least 30 minutes; removing the schistosomes from the saline extracting solution; and, collecting the remaining solution containing said live worm-derived antigen-containing extractant. ### Claim 2 (depends on 1) The antigen containing extractant of claim 1, wherein the saline solution for said perfusion, said rinsing, and said extracting comprise sodium phosphate buffered sodium chloride. ### Claim 3 (depends on 2) The antigen-containing extractant of claim 2, wherein the perfusion solution and extracting solution comprise 0.15 M NaCl buffered to pH 6.8 by sodium phosphate. ### Claim 4 (depends on 3) The antigen-containing extractant of claim 3, wherein the schistosomes are removed from the saline extracting solution by centrifuging the extracting solution, and the remaining solution comprises the supernatant from said centrifuging step. ### Claim 5 (depends on 3) The antigen-containing extractant as set forth in claim 3, wherein the schistosomes are removed from the saline extraction solution by filtering said extraction solution through an exclusion pore filter having a pore size which retains solution components having molecular weight greater than about 50,000, and the remaining solution comprises the material retained on said filter. ### Claim 6 (depends on 3) The antigen-containing extractant as set forth in claim 3, wherein the schistosomes are removed from the saline extraction solution by filtering said extraction solution through an exclusion pore filter having a pore size which retains solution components having molecular weight greater than about 100,000, and the remaining solution comprises the material retained on said filter. ### Claim 7 (depends on 1) The antigen-containing extractant of claim 1 wherein the step of collecting comprises collecting the schistosomes on a sieve whose openings will not allow the worms to pass through and wherein the step of rinsing comprises slowly pouring or spraying the collected schistosomes from one to three times with a total volume of saline solution of from about five to about 30 ml per 100 worms. ### Claim 8 (depends on 7) The antigen-containing extractant of claim 7 wherein, in the step of extracting, the rinsed adult schistosomes are contacted with from about 1.0 ml to about 20 ml of saline extracting solution per gram of schistosomes. ### Claim 9 A vaccine for immunizing mammals against infection by schistosomes comprising an antigenic material containing saline extract of live adult schistosomes obtained by the steps of: perfusing fresh, living adult schistosome worms from a host with a buffered saline solution; rinsing the perfused worms with from one to three low volume buffered saline solutions to remove only blood and non-worm blood solids, said rinsing not washing away any significant amount of antigenic material; contacting the rinsed worms with buffered saline extracting solution for a period of at least 30 minutes to obtain a saline extract antigenic material from said worms; centrifuging said saline extract for a period of time sufficient to remove substantially all of the worm bodies and other suspended solid particles from said saline extract, and recovering the supernatant which is at least substantially, completely free from suspended solids; eluting the supernatant through a gel partition chromatography column capable of separating high molecular weight components, wherein the eluant is buffered saline solution; and, recovering the eluted fractions containing the antigenic material. ### Claim 10 (depends on 9) The vaccine, according to claim 9, wherein before said centrifuging step, the low molecular weight components of said saline extract are removed by filtering the saline extract through an exclusion pore filter having a pore size which retains solution components having molecular weight greater than about 50,000, and recovering the saline extract retained on said filter for further processing. ### Claim 11 (depends on 10) The vaccine according to claim 10, wherein said exclusion pore filter has a pore size which retains solution components having molecular weight greater than about 100,000. ### Claim 12 (depends on 9) The vaccine according to claim 9 wherein the gel partition chromatography column is one which has an exclusion limit in the range of from about 100,000 to about 400,000, and the antigenic material is recovered in the first and second eluted fractions. ### Claim 13 (depends on 12) The vaccine according to claim 12 wherein the antigenic material recovered in the first eluted fraction comprises a first glycoprotein having a molecular weight in the range of from about 200,000 to less than about 400,000 and a second glycoprotein having a molecular weight in the range of from about 100,000 to about 200,000, wherein the glyco moiety of said glycoproteins is a mannose sugar moiety; and wherein the antigenic material recovered in the second eluted fraction comprises a protein having a molecular weight in the range of from about 40,000 to about 60,000, wherein said molecular weights are determined by gel filtration. ### Claim 14 A method of extracting antigenic material from live adult schistosomes comprising the steps of: collecting fresh, living adult schistosomes from an infected host without destroying the schistosomes; rinsing the fresh, living adult schistosomes with saline solution to remove only blood and non-worm solids but without removing worm by-products; storing the collected adult schistosomes in a saline extracting solution for a period of at least about 30 minutes; removing the schistosomes from the saline extracting solution; and, collecting the remaining solution containing live worm-derived antigenic substance. ### Claim 15 (depends on 14) The method as set forth in claim 14 wherein the saline extracting solution consists essentially of a 0.15 M sodium chloride solution with a sodium phosphate buffer at pH 6.8. ### Claim 16 (depends on 15) The method as set forth in claim 15 wherein the step of removing comprises centrifuging the extracting solution, and collecting the supernatant as the remaining solution. ### Claim 17 (depends on 15) The method as set forth in claim 15 wherein the step of removing comprises filtering the extracting solution through a filter having an exclusion pore size which retains solution components having a molecular weight greater than about 50,000, and retaining the high molecular weight solution. ### Claim 18 (depends on 15) The method as set forth in claim 15 wherein the step of removing comprises filtering the extracting solution through a filter having an exclusion pore size which retains solution components having a molecular weight greater than about 100,000, and retaining the high molecular weight solution. ### Claim 19 (depends on 17, 18) The method as set forth in claim 17 or 18 which further comprises centrifuging the remaining solution and collecting the supernatant as the remaining solution. ### Claim 20 (depends on 16, 17) The method as set forth in claim 16 or 17 which further comprises purifying the remaining solution by passing the remaining solution through a gel partition chromatography column capable of separating high molecular weight components using buffered saline solution as the eluant, and recovering the first and second eluted fractions containing said antigenic material. ### Claim 21 (depends on 9) A method of immunizing a mammal against infection by schistosomes comprising administering to said mammal an effective dose of the antigen-containing vaccine of claim 9 to induce an immunological reaction. ### Claim 22 (depends on 21) The method of immunizing a mammal against infection by schistosomes as set forth in claim 21 further including the steps of periodically administering additional amounts of said vaccine as a booster to reinforce the immune reaction in the mammal. ### Claim 23 (depends on 21) The method according to claim 21 wherein said mammal is a human.