Reagent for assaying endotoxin
First Claim
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1. A reagent for assaying an endotoxin comprising limulus amebocyte lysate and an antibody to (1 →
- 3)-β
-D-glucan sensitive factor, wherein said antibody specifically binds to and neutralizes (1 →
3)-β
-D-glucan sensitive factor.
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Abstract
The present invention provides a reagent for assaying an endotoxin which comprises limulus amoebocyte lysate and an antibody to (1 → 3)-β-D-glucan sensitive factor or a reagent which comprises a lysate substantially free from (1 → 3)-β-D-glucan sensitive factor. The reagent of the present invention makes it possible to assay an endotoxin originating from gram-negative bacteria contained in a biological sample such as blood, urine and cerebrospinal fluid at an extremely high sensitivity without being affected by (1 → 3)-β-D-glucan.
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Citations
11 Claims
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1. A reagent for assaying an endotoxin comprising limulus amebocyte lysate and an antibody to (1 →
- 3)-β
-D-glucan sensitive factor, wherein said antibody specifically binds to and neutralizes (1 →
3)-β
-D-glucan sensitive factor. - View Dependent Claims (2, 3, 8)
- 3)-β
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4. A reagent for assaying an endotoxin comprising limulus amebocyte lysate, an antibody to (1 →
- 3)-β
-D-glucan sensitive factor, wherein said antibody specifically binds to and neutralizes (1 →
3)-β
-D-glucan sensitive factor, and a synthetic peptide substrate for clotting enzyme having an amino acid sequence in common with the hydrolysis sites of coagulogen. - View Dependent Claims (5, 6)
- 3)-β
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7. A reagent for assaying an endotoxin comprising a limulus amebocyte lysate substantially free from (1 →
- 3)-β
-D-glucan sensitive factor, obtained by method (I) or (II), wherein said method (I) comprises the steps of;(a) admixing limulus amebocyte lysate with a carrier onto which an antibody to (1 →
3)-β
-D-glucan sensitive factor has been immobilized, wherein said antibody specifically binds to and neutralizes (1 →
3)-β
-D-glucansensitive factor, and (b) removing the carrier from the lysate so as to obtain said reagent; wherein said method (II) comprises the steps of; (a) applying limulus amebocyte lysate to a column which has previously been packed with a carrier onto which an antibody to (1 →
3)-β
-D-glucan sensitive factor has been immobilized, wherein said antibody specifically binds to and neutralizes (1 →
3)-β
-D-glucan sensitive factor, and(b) collecting a fraction of the lysate which passes through the column so as to obtain said reagent. - View Dependent Claims (9, 10)
- 3)-β
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11. A method of assaying for an endotoxin in a sample comprising:
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(a) admixing limulus amebocyte lysate with an antibody to (1 →
3)-β
-D-glucan sensitive factor, wherein said antibody specifically binds to and neutralizes (1 →
3)-β
-D-glucan sensitive factor in the lysate,(b) combining the admixed lysate with a sample under conditions whereby endotoxin in the sample, acting via an endotoxin-sensitive factor-dependent cascade, activates clotting enzyme in the lysate, (c) contacting the activated clotting enzyme with a synthetic peptide substrate for clotting enzyme having an amino acid sequence in common with the hydrolysis sites of coagulogen, and (d) measuring enzymatic activity of the activated clotting enzyme on coagulogen or on the synthetic peptide substrate as an indication of endotoxin in the sample.
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Specification