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US 10,982,202 B2Granted
Recombinant factor C and method for producing the same, and method for measuring endotoxin
AssigneeSeikagaku Corporation
InventorsHikaru Mizumura, Toshio Oda, Shun-ichiro Kawabata
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Abstract
To provide a method for producing a horseshoe crab recombinant Factor C. The horseshoe crab recombinant Factor C is produced through expression thereof by use of mammalian cells such as CHO DG44 and HEK293 as host cells.
Claims13
A method for imparting a salt ion resistance for a horseshoe crab Factor C, the method comprising:
- recombinantly expressing and secreting the horseshoe crab Factor C into a culture broth or a culture supernatant using a mammalian cell as a host cell, and
- recovering the recombinant horseshoe crab Factor C secreted from the mammalian cell,
- wherein the horseshoe crab Factor C recombinantly expressed and secreted using the mammalian cell contains (α-2,3)-linked terminal sialic acid in a greater amount, as compared with a corresponding horseshoe crab Factor C expressed using a Sf9 insect cell as a host cell,
- wherein the mammalian cell is a Chinese hamster ovary (CHO) cell or a human embryonic kidney (HEK) cell, and
- wherein the horseshoe crab Factor C recombinantly expressed and secreted into a culture broth or a culture supernatant using the mammalian cell is a protein selected from the following (A), (B), (C), or (D):
- (A) a protein comprising the amino acid sequence of Factor C of Tachypleus tridentatus, wherein the protein has no peptide added to the N-terminus of Factor C of Tachypleus tridentatus, and wherein the protein has activity of Factor C;
- (B) a protein comprising the amino acid sequence of Factor C of Limulus polyphemus, wherein the protein has no peptide added to the N-terminus of Factor C of Limulus polyphemus, and wherein the protein has activity of Factor C;
- (C) a protein comprising the amino acid sequence of Factor C of Carcinoscorpius rotundicauda, wherein the protein has no peptide added to the N-terminus of Factor C of Carcinoscorpius rotundicauda, and wherein the protein has activity of Factor C;
- (D) a protein comprising the amino acid sequence of Factor C of Tachypleus gigas, wherein the protein has no peptide added to the N-terminus of Factor C of Tachypleus gigas, and wherein the protein has activity of Factor C.
6 more elements
A method for producing an endotoxin assay agent comprising a horseshoe crab Factor C, the method comprising:
- recombinantly expressing and secreting the horseshoe crab Factor C into a culture broth or a culture supernatant using a mammalian cell as a host cell,
- recovering the recombinant horseshoe crab Factor C secreted from the mammalian cell, and
- producing an endotoxin assay agent comprising the recombinant horseshoe crab Factor C,
- wherein the horseshoe crab Factor C recombinantly expressed and secreted using the mammalian cell contains (α-2,3)-linked terminal sialic acid in a greater amount, as compared with a corresponding horseshoe crab Factor C expressed using a Sf9 insect cell as a host cell,
- wherein the mammalian cell is a Chinese hamster ovary (CHO) cell or a human embryonic kidney (HEK) cell, and
- wherein the horseshoe crab Factor C recombinantly expressed and secreted into a culture broth or a culture supernatant using the mammalian cell is a protein selected from the following (A), (B), (C), or (D):
- (A) a protein comprising the amino acid sequence of Factor C of Tachypleus tridentatus, wherein the protein has no peptide added to the N-terminus of Factor C of Tachypleus tridentatus, and wherein the protein has activity of Factor C;
- (B) a protein comprising the amino acid sequence of Factor C of Limulus polyphemus, wherein the protein has no peptide added to the N-terminus of Factor C of Limulus polyphemus, and wherein the protein has activity of Factor C;
- (C) a protein comprising the amino acid sequence of Factor C of Carcinoscorpius rotundicauda, wherein the protein has no peptide added to the N-terminus of Factor C of Carcinoscorpius rotundicauda, and wherein the protein has activity of Factor C;
- (D) a protein comprising the amino acid sequence of Factor C of Tachypleus gigas, wherein the protein has no peptide added to the N-terminus of Factor C of Tachypleus gigas, and wherein the protein has activity of Factor C.
7 more elements
Dependencies are inferred from the claim text.