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Mitochondria creatinekinase antibody

A creatine kinase and mitochondrial technology, applied in the field of clinical examination, can solve the problems of lack of anti-umCK antibody and difficulty in distinguishing

Inactive Publication Date: 2003-11-12
SYSMEX CORP
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the determination of mCK generally adopts the agarose gel electrophoresis method. Since the electrophoresis of umCK and smCK is basically the same, the two are reported indiscriminately.
Although an antibody with anti-umCK activity has been reported (JP-A-2002-270 Publication), there is no report on an anti-umCK antibody, so it is difficult to distinguish and analyze the two

Method used

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  • Mitochondria creatinekinase antibody
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  • Mitochondria creatinekinase antibody

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0082] Example 1: Production of hybrid cell tumors producing monoclonal antibodies (MoAb)

[0083] (1) Modulation of immunogen (antigen)

[0084] The present invention uses human umCK as an antigen (Lot No. ETH010122 manufactured by the Institute of Cell Biology).

[0085] (2) Place the immunized 5-8 week-old inbred BALB / c female mice in the animal room (23±1°C, temperature 70%), feed them with standard granules, and give them water arbitrarily.

[0086] (3) The refined product prepared in the above (1) with human umCK as the antigen was prepared with PBS to a concentration of 100μg / 0.5mL and an equal amount (0.5mL) of Freund's complete adjuvant (frend's complete adjubant: manufactured by Difco) was mixed and emulsified. Inject 200 μL of this emulsified antigen into the abdominal cavity of 5-week-old female BALB / C mice, and then inject 100 μg / mL of the above-mentioned antigen prepared with Ribi adjuvant into each mouse every 2 weeks. Inject 20 μg, four times in total. Then...

experiment example 1

[0101] Experimental example 1: Regarding the specificity of enzyme repression of culture supernatant

[0102] With respect to the culture supernatant of the hybrid cell tumor-producing cells in the 7 acupoints screened above, inhibition of each enzyme activity was investigated using human umCK, human smCK, human CK-MM, human CK-BB, or human CK-MB, and the effect on the activity of each enzyme was confirmed. Specificity of human CK isozymes. For verification, a MoAb of mCKI-578 (JP-A 2002-270 publication) produced by a hybrid cell tumor commissioned under the commission number FERM BP-7133 was also investigated.

[0103] The result is as figure 1 shown. The supernatants of pure lines UI-178, UI-281, UI-956, UI-1111, UI-1125, UI-1881 and UI-1299 can inhibit more than 82% of umCK enzyme activity, but for smCK, CK- The enzymatic activities of MM, CK-BB, and CK-MB were not suppressed at all. Especially the supernatant of UI-1881 could repress 94% of umCK enzyme activity. On th...

experiment example 2

[0104] Experimental Example 2: Establishment of a MoAb Hybrid Cell Tumor Line (Selection Establishment)

[0105] The above-mentioned hybrid cell tumors in the 7 cavities obtained by screening were screened by the limiting dilution method, and one pure line was selected that could stably produce hybrid cell tumors showing inhibition of mCK enzyme activity. Using this cell as the establishment strain, we entrusted the processing to the Institute of Bioengineering and Industrial Technology, Institute of Industrial Technology, Ministry of International Trade and Industry, under the consignment number FERM P-18760.

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Abstract

The present invention refers to an anti-mCK antibody characterized by inhibiting an enzyme activity of ubiquitous mCK (umCK) among creatine kinase isozymes localized in mitochondria (mCK).

Description

technical field [0001] The invention relates to the field of clinical examination, in particular to a quantitative method for measuring creatine kinase (hereinafter referred to as CK) isoenzyme, an antibody used in the determination, a reagent for determination and a matching kit for the reagent for determination. Background technique [0002] There are four proteins in human CK that cause gene mutation. Two proteins (a localized muscle type (M type) and a brain type (B type)) are produced from the cytoplasm and the other from the mitochondria. The CK isoenzymes produced in the cytoplasm are composed of diploids formed by the combination of the M type and the B type, and are divided into 3 types: CK-MM, CK-MB, and CK-BB. Mitochondrial CK has two types, although it is composed of octoploids and is very stable, but in the presence of similar substance complexes in the migratory state of creatine Mg, ADP and nitrate, the dissociation will be accelerated to 2 times in a few min...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/573C07K16/40C12N5/10C12N5/20C12P21/08G01N33/577
CPCY10S435/975C07K16/40
Inventor 白波瀨泰史梶田忠宏岸浩司山下和昭
Owner SYSMEX CORP