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Detecting human antibodies in non-human serum

An antibody and non-human technology, applied in measuring devices, biological testing, material inspection products, etc., can solve problems such as difficulty in obtaining

Inactive Publication Date: 2008-02-27
GENENTECH INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Such target-specific molecules that bind anti-CD20 antibodies are not readily available

Method used

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  • Detecting human antibodies in non-human serum
  • Detecting human antibodies in non-human serum
  • Detecting human antibodies in non-human serum

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0121] Example 1: Pre-absorption to reduce background / variation

[0122] 1. Pre-absorption to remove cross-reactivity of rhesus monkey IgG

[0123] A variety of capture agents are screened for use in the methods disclosed in the present invention. Sheep anti-human IgG heavy chain (H) and light chain (L) (catalog number CUS 1684) absorbed with rhesus monkey serum were purchased from The Binding Site (San Diego, CA) and showed promising potential. However, as discussed below and shown in Table 3, the use of this capture agent produces a high and variable background. Therefore, in an attempt to reduce background and background variation, purified rhesus IgG obtained from problematic individual monkey (high background) serum was used to further absorb sheep anti-human IgG (H+L) absorbed by monkey serum to further Remove any IgG binding components in the capture reagent.

[0124] First, use HiTrap protein G column (Pharmacia) to purify cynomolgus monkey IgG following the procedure reco...

Embodiment 2

[0138] Example 2: BGG (bovine gamma globulin) in blocking buffer

[0139] Analyze the use of BGG in blocking buffer, sample diluent, and detector diluent to determine whether the background level and background variation can be improved. Sheep anti-human IgG heavy chain (H) and light chain (L) capture reagent (catalog number CUS1684) and sheep anti-human IgG (H+L) HRP conjugate detector (catalog number CUS 1684.H) were purchased from The Binding Site (San Diego, CA). The humanized mAb can be produced by known methods. rhuMAb2H7 was produced as described in WO 04 / 056312. Herceptin, Xolair, Avastin TM And Raptiva(R) were produced according to the procedures described in Carter et al., PNAS 89: 4285-4289 (1992), U.S. Patent No. 6,172,213, Presta et al., Cancer Research 57: 4593-4599 (1997) and U.S. Patent No. 6,037,454, respectively. Goat anti-human IgG (H+L) horseradish peroxidase (HRP) (detection agent) was purchased from American Qualex (San Clemente, CA). Individual rhesus monkey...

Embodiment 3

[0182] Example 3: Comparison of bridged ELISA and direct ELISA

[0183] The assay described in Example 2 is a direct ELISA protocol in which the detection agent is different from the capture agent. Bridged ELISA in which the detection agent and the capture agent contain the same reagent (such as the same antibody) are known to produce reduced non-specific background in some assays.

[0184] The bridge format assay system is tested to compare the background and background variation obtained with the direct ELISA process.

[0185] Comparison of the background and variation of rhesus monkey serum with different detection agents was performed in two diluents. Goat anti-human IgG (H+L)·HRP and sheep anti-human IgG (H+L)·HRP absorbed by monkey serum are used in direct and bridged formats respectively. Sheep anti-human IgG (H+L) is used as a capture agent. Buffer D is used to seal the plate and dilute the sample. The ELISA assay was performed as described in Example 2. RhuMAb2H7 samples ...

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Abstract

The present invention provides methods for the quantification of human, human chimeric, humanized antibodies or fragments thereof without the use of target specific molecules. More specifically, the present invention relates to a quantitative assay comprising the step of blocking the non-specific binding sites of a capture agent with a blocking buffer comprising a non-human mammalian globulin such as bovine gamma globulin (BGG).

Description

[0001] This application is for applicants from all designated countries except the United States, the United States state-owned company Genentech (Genentech, Inc.) and Chinese citizens Jihong Yang and U.S. citizens Valerie Elizabeth Quarmby, as a PCT international patent application in December 2005 Filed on December 30, and claim the priority of U.S. Provisional Patent Application No. 60 / 640,948 filed on December 31, 2004. Background of the invention [0002] The potential therapeutic applications of antibodies, including humanized monoclonal antibodies, in humans have been extensively studied. In the early development of therapeutic antibodies, the effectiveness and safety of molecules were studied. Generally, such research involves the administration of antibodies to non-human species such as non-human primates. Analyze the presence and concentration of the antibody of interest in body fluids, such as serum, obtained from non-human species. To perform this analysis, a sensitive ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/68
CPCG01N33/54393G01N33/6854G01N33/54333
Inventor 杨蓟红瓦莱丽·E·夸姆比
Owner GENENTECH INC
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