Borrelia antigen

A composition which binds with antibodies elicited by B. burgdorferi, said composition comprising an effective amount of fraction B for antibody binding, and a carrier or vehicle, wherein said fraction B is obtained by the following method comprising the steps:

• a) lysing B. burgdorferi spirochaete cells with a detergent so as to release outer membranae components from the cells but not denature said outer membrane components,
• b) subsequently subjecting the lysed cells to centrifugation resulting in a first pellet comprising cell wall and flagellar components and a first supernatant comprising outer membrane components,
• c) incubating the first supernatant from step b) under conditions sufficient to precipitate proteins of fraction E of the first supernatant followed by centrifugation so as to obtain a second pellet comprising fraction E and a second supernatant,
• d) subjecting the second supernatant from step c) to filtration and dialyzing the supernatant against an aqueous medium with a low ionic strength so as to substantially remove the detergent and allow B. burgdorferi derived cell components to precipitate in the dialysis bag,
• e) centrifuging the contents of the dialysis bag so as to obtain a third pellet comprising fraction B and a third supernatant comprising fraction C,
• the fraction B containing lipids and being
• substantially free from cell wall and flagellar components of B. burgdorferi,
• substantially free from detergent, and
• substantially free from sodium dodecyl sulphate.

A diagnostic composition for the detection of B. burgdorferi antibodies in a sample, which comprises fraction B of B. burgdorferi wherein said fraction B is obtained by the following method comprising the steps:

• a) lysing B. burgdorferi spirochaete cells with a detergent so as to release outer membranae components from the cells but not denature said outer membrane components,
• b) subsequently subjecting the lysed cells to centrifugation resulting in a first pellet comprising cell wall and flagellar components and a first supernatant comprising outer membrane components,
• c) incubating the first supernatant from step b) under conditions sufficient to precipitate proteins of fraction E of the first supernatant followed by centrifugation so as to obtain a second pellet comprising fraction E and a second supernatant,
• d) subjecting the second supernatant from step c) to filtration and dialyzing the supernatant against an aqueous medium with a low ionic strength so as to substantially remove the detergent and allow B. burgdorferi derived cell components to precipitate in the dialysis bag,
• e) centrifuging the contents of the dialysis bag so as to obtain a third pellet comprising fraction B and a third supernatant comprising fraction C,
• the fraction B containing lipids and being
• substantially free from cell wall and flagellar components of B. burgdorferi,
• substantially free from detergent, and
• substantially free from sodium dodecyl sulphate.

A method of preparing fraction B of B. burgdorferi which fraction B contains lipids, is substantially free from cell wall and flagellar components of B burgdorferi, is substantially free from detergent, and, is substantially free from sodium dodecyl sulphate, said method consisting essentially of the following sequential steps:

• a) lysing B. burgdorferi spirochaete cells with a detergent so as to release outer membrane components from the cells but not denature said outer membrane components,
• b) subsequently subjecting the lysed cells to centrifugation resulting in a first pellet comprising cell wall and flagellar components and a first supernatant comprising outer membrane components,
• c) separating said first supernatant from said first pellet,
• d) subsequently incubating the separated first supernatant from step c.) under conditions sufficient to precipitate proteins of fraction E of the first supernatant followed by centrifugation so as to obtain a second pellet comprising fraction E and a second supernatant,
• e) separating said second supernatant from said second pellet,
• f) subsequently subjecting the separated second supernatant from step e) to filtration to remove insoluble particles from said separated second supernatant,
• g) subsequently dialyzing the filtered second supernatant against an aqueous medium with a low ionic strength so as to substantially remove the detergent and allow B. burgdorferi derived cell components to precipitate in the dialysis bag,
• h) subsequently centrifuging the contents of the dialysis bag so as to obtain a third pellet comprising fraction B and a third supernatant comprising fraction C, and
• i) subsequently separating said third pellet from said third supernatant.

A method of preparing fraction B of B. burgdorferi which fraction B contains lipids, is substantially free from cell wall and flagellar components of B. burgdorferi, is substantially free from detergent, and, is substantially free from sodium dodecyl sulphate, said method, consisting essentially of the following sequential steps:

• a) lysing B. burgdorferi spirochaete cells with a detergent so as to release outer membrane components from the cells but not denature said outer membrane components,
• b) subsequently subjecting the lysed cells to centrifugation resulting in a first pellet comprising cell wall and flagellar components and a first supernatant comprising outer membrane components,
• c) separating said first supernatant from said first pellet,
• d) subsequently incubating the separated first supernatant from step c) under conditions sufficient to precipitate proteins of fraction E of the first supernatant followed by centrifugation so as to obtain a second pellet comprising fraction E and a second supernatant,
• e) separating said second supernatant from said second pellet,
• f) subsequently subjecting the separated second supernatant from step e) to filtration to remove insoluble particles from said separated second supernatant,
• g) subsequently subjecting the filtered second supernatant to ultrafiltration so as to substantially remove the detergent and allow B burgdorferi derived cell components to precipitate in the retentate resulting from the ultrafiltration,
• h) subsequently centrifuging the retentate so as to obtain a third pellet comprising fraction B and a third supernatant comprising fraction C, and
• i) subsequently separating said third pellet from said third supernatant.