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Human Creatine Kinase MM / CKMM ELISA Kit

SKU:
HUFI00465
Product Type:
ELISA Kit
Size:
96 Assays
Uniprot:
P06732
Sensitivity:
0.938ng/ml
Range:
1.563-100ng/ml
ELISA Type:
Sandwich
Synonyms:
CKM, CKMM, M-CK, creatine kinase M-type, Creatine kinase M chain, creatine kinase, muscle, creatine kinase-M
Reactivity:
Human
Research Area:
Cell Biology
$719
Frequently bought together:

Description

Human Creatine Kinase MM / CKMM ELISA Kit

Creatine Kinase MM / CKMM is a cytoplasmic enzyme involved in energy homeostasis and is an important serum marker for myocardial infarction. Creatine Kinase MM / CKMM reversibly catalyzes the transfer of phosphate between ATP and various phosphogens such as creatine phosphate. Creatine Kinase MM / CKMM acts as a homodimer in striated muscle, and as a heterodimer with a similar brain isozyme in heart. Diseases associated with Creatine Kinase MM / CKMM include Myotonic Dystrophy and Mcleod Syndrome.

system_update_alt Datasheet system_update_alt MSDS

Key Features

Save Time Pre-coated 96 well plate
Quick Start Kit includes all necessary reagents
Publication Ready Reproducible and reliable results

Overview

Product Name:

Human Creatine Kinase MM / CKMM ELISA Kit

Product Code:

HUFI00465

Size:

96 Assays

Alias:

CKM, CKMM, M-CK, creatine kinase M-type, Creatine kinase M chain, creatine kinase, muscle, creatine kinase-M

Detection Method:

Sandwich ELISA, Double Antibody

Application:

This immunoassay kit allows for the in vitro quantitative determination of Human CKM concentrations in serum plasma and other biological fluids.

Sensitivity:

0.938ng/ml

Range:

1.563-100ng/ml

Storage:

4°C for 6 months

Note:

For Research Use Only

Additional Information

Recovery:

Matrices listed below were spiked with certain level of Human CKM and the recovery rates were calculated by comparing the measured value to the expected amount of Human CKM in samples.

Matrix

Recovery Range (%)

Average (%)

serum (n=5)

87-103

98

EDTA plasma (n=5)

86-100

91

UFH plasma (n=5)

86-99

93

Linearity:

The linearity of the kit was assayed by testing samples spiked with appropriate concentration of Human CKM and their serial dilutions. The results were demonstrated by the percentage of calculated concentration to the expected.

Sample

1:2

1:4

1:8

Serum (n=5)

90-105%

90-104%

85-98%

EDTA plasma (n=5)

86-101%

82-99%

87-99%

UFH Plasma (n=5)

83-91%

81-98%

82-85%

CV(%):

Intra-Assay: CV<8%
Inter-Assay: CV<10%

Kit Components

Component Quantity Storage

ELISA Microplate (Dismountable)

8x12 strips

4°C for 6 months

Lyophilized Standard

2

4°C/ -20°C

Sample/Standard Dlution Buffer

20ml

4°C

Biotin-labeled Antibody (Concentrated)

120ul

4°C (Protection from light)

Antibody Dilution Buffer

10ml

4°C

HRP-Streptavidin Conjugate (SABC)

120ul

4°C (Protect from light)

SABC Dilution Buffer

10ml

4°C

TMB Substrate

10ml

4°C (Protection from light)

Stop Solution

10ml

4°C

Wash Buffer (25X)

30ml

4°C

Plate Sealer

5

-

Other materials required:

  • Microplate reader with 450 nm wavelength filter
  • Multichannel Pipette, Pipette, microcentrifuge tubes and disposable pipette tips
  • Incubator
  • Deionized or distilled water
  • Absorbent paper
  • Buffer resevoir

Protein Information

Uniprot:

UniProt Protein Function:

CKM: Reversibly catalyzes the transfer of phosphate between ATP and various phosphogens (e.g. creatine phosphate). Creatine kinase isoenzymes play a central role in energy transduction in tissues with large, fluctuating energy demands, such as skeletal muscle, heart, brain and spermatozoa. Belongs to the ATP:guanido phosphotransferase family.

UniProt Code:

NCBI GenInfo Identifier:

NCBI Gene ID:

NCBI Accession:

UniProt Related Accession:

Molecular Weight:

Calculated: 40kDaObserved: 43kDa

Protocol

*Note: Protocols are specific to each batch/lot. For the exact instructions please follow the protocol included in your kit.

Before adding to wells, equilibrate the SABC working solution and TMB substrate for at least 30 min at 37°C. When diluting samples and reagents, they must be mixed completely and evenly. It is recommended to plot a standard curve for each test.

Step Procedure

1.

Set standard, test sample and control (zero) wells on the pre-coated plate respectively, and then, record their positions. It is recommended to measure each standard and sample in duplicate. Wash plate 2 times before adding standard, sample and control (zero) wells!

2.

Aliquot 0.1ml standard solutions into the standard wells.

3.

Add 0.1 ml of Sample / Standard dilution buffer into the control (zero) well.

4.

Add 0.1 ml of properly diluted sample ( Human serum, plasma, tissue homogenates and other biological fluids.) into test sample wells.

5.

Seal the plate with a cover and incubate at 37 °C for 90 min.

6.

Remove the cover and discard the plate content, clap the plate on the absorbent filter papers or other absorbent material. Do NOT let the wells completely dry at any time. Wash plate X2.

7.

Add 0.1 ml of Biotin- detection antibody working solution into the above wells (standard, test sample & zero wells). Add the solution at the bottom of each well without touching the side wall.

8.

Seal the plate with a cover and incubate at 37°C for 60 min.

9.

Remove the cover, and wash plate 3 times with Wash buffer. Let wash buffer rest in wells for 1 min between each wash.

10.

Add 0.1 ml of SABC working solution into each well, cover the plate and incubate at 37°C for 30 min.

11.

Remove the cover and wash plate 5 times with Wash buffer, and each time let the wash buffer stay in the wells for 1-2 min.

12.

Add 90 µl of TMB substrate into each well, cover the plate and incubate at 37°C in dark within 10-20 min. (Note: This incubation time is for reference use only, the optimal time should be determined by end user.) And the shades of blue can be seen in the first 3-4 wells (with most concentrated standard solutions), the other wells show no obvious color.

13.

Add 50 µl of Stop solution into each well and mix thoroughly. The color changes into yellow immediately.

14.

Read the O.D. absorbance at 450 nm in a microplate reader immediately after adding the stop solution.

Sample Type

When carrying out an ELISA assay it is important to prepare your samples in order to achieve the best possible results. Below we have a list of procedures for the preparation of samples for different sample types.

Sample Type Protocol

Serum

If using serum separator tubes, allow samples to clot for 30 minutes at room temperature. Centrifuge for 10 minutes at 1,000x g. Collect the serum fraction and assay promptly or aliquot and store the samples at -80°C. Avoid multiple freeze-thaw cycles.

If serum separator tubes are not being used, allow samples to clot overnight at 2-8°C. Centrifuge for 10 minutes at 1,000x g. Remove serum and assay promptly or aliquot and store the samples at -80°C. Avoid multiple freeze-thaw cycles.

Plasma

Collect plasma using EDTA or heparin as an anticoagulant. Centrifuge samples at 4°C for 15 mins at 1000 × g within 30 mins of collection. Collect the plasma fraction and assay promptly or aliquot and store the samples at -80°C. Avoid multiple freeze-thaw cycles. Note: Over haemolysed samples are not suitable for use with this kit.

Urine & Cerebrospinal Fluid

Collect the urine (mid-stream) in a sterile container, centrifuge for 20 mins at 2000-3000 rpm. Remove supernatant and assay immediately. If any precipitation is detected, repeat the centrifugation step. A similar protocol can be used for cerebrospinal fluid.

Cell culture supernatant

Collect the cell culture media by pipette, followed by centrifugation at 4°C for 20 mins at 1500 rpm. Collect the clear supernatant and assay immediately.

Cell lysates

Solubilize cells in lysis buffer and allow to sit on ice for 30 minutes. Centrifuge tubes at 14,000 x g for 5 minutes to remove insoluble material. Aliquot the supernatant into a new tube and discard the remaining whole cell extract. Quantify total protein concentration using a total protein assay. Assay immediately or aliquot and store at ≤ -20 °C.

Tissue homogenates

The preparation of tissue homogenates will vary depending upon tissue type. Rinse tissue with 1X PBS to remove excess blood & homogenize in 20ml of 1X PBS (including protease inhibitors) and store overnight at ≤ -20°C. Two freeze-thaw cycles are required to break the cell membranes. To further disrupt the cell membranes you can sonicate the samples. Centrifuge homogenates for 5 mins at 5000xg. Remove the supernatant and assay immediately or aliquot and store at -20°C or -80°C.

Tissue lysates

Rinse tissue with PBS, cut into 1-2 mm pieces, and homogenize with a tissue homogenizer in PBS. Add an equal volume of RIPA buffer containing protease inhibitors and lyse tissues at room temperature for 30 minutes with gentle agitation. Centrifuge to remove debris. Quantify total protein concentration using a total protein assay. Assay immediately or aliquot and store at ≤ -20 °C

Breast Milk

Collect milk samples and centrifuge at 10,000 x g for 60 min at 4°C. Aliquot the supernatant and assay. For long term use, store samples at -80°C. Minimize freeze/thaw cycles.

CKMM Background

Creatine Kinase MM, also known as CK-MM or creatine kinase muscle-type, is one of the three isoenzymes of creatine kinase. The other two isoenzymes are creatine kinase BB (CK-BB) found in the brain and creatine kinase MB (CK-MB) found mainly in the heart.

Function of CKMM

The primary function of Creatine Kinase MM revolves around energy metabolism in muscle tissues. Within cells, particularly in skeletal muscles, CK-MM catalyzes the reversible transfer of a phosphate group between creatine and ADP, converting them into creatine phosphate and ATP, respectively. This reaction is fundamental for cellular energy homeostasis, as ATP is the primary energy currency for numerous cellular processes.

During periods of heightened energy demand, such as intense physical activity or muscle contraction, the CK-MM reaction facilitates the rapid replenishment of ATP stores, ensuring sustained energy supply to meet the cellular needs. Consequently, CK-MM's critical role in energy metabolism makes it a key player in the functioning of skeletal muscles.

Clinical Significance of CK-MM Levels

CK-MM levels in the blood can be measured and are often used as a marker for muscle damage or injury. When there is damage to muscle cells due to trauma, strenuous exercise, or certain medical conditions, CK-MM is released into the bloodstream, leading to an increase in its concentration. Medical professionals can use CK-MM levels, along with other clinical information and tests, to diagnose and monitor conditions such as muscle disorders, muscular dystrophy, muscle inflammation (myositis), and rhabdomyolysis (a severe breakdown of muscle tissue). Elevated CK-MM levels can indicate muscle injury or disease, while decreased levels may be seen in certain muscle-related disorders.

CKMM FAQs

What is the CKMM ELISA Kit?

The CKMM ELISA Kit is an advanced assay designed to accurately quantify Human Creatine Kinase MM levels in biological samples.

What are the advantages of using the CKMM ELISA Kit?

The CKMM ELISA Kit offers high sensitivity, specificity for Human CKMM, an easy-to-use protocol, quantitative results, compatibility with various samples, and time-efficiency for research screening.

Where can I find more information about the CKMM ELISA Kit?

For more detailed information about the CKMM ELISA Kit, including technical specifications, performance characteristics, and ordering details, please refer to the product brochure or contact our customer support team. We are here to assist you with any inquiries you may have.