‘Acrolein’ ELISA Kit
ELISA kit for ACR-Lys adduct (FDP-Lys)
Catalog #KACR-200E
What is ACROLEIN?
Acrolein(CH2=CH-CHO; acrolein, ACR) is a chemical substance formed not only by the combustion of petroleum, charcoal, lumber and plastic but also by cigarette smoke, exhaust gas, heating of oils and fats and is known as a strong cytotoxic substance. In recent years it has been revealed by Uchida et al., of Nagoya University that ACR can also be produced by hyperoxidation of lipid and its presence as a secondary product resulting from hyperoxidation of lipid in vivo was identified, and further, ACR's effects on the living body has become a focus of attention. This product is an immunological measuring kit, using a monoclonal antibody which reacts specifically to an ACR-lys added compound, for example, formyldehydro- piperidino- lysine structure produced by the addition of ACR to protein lysine residues and utilizing the competitive method dedicated to measuring ACR-lys adduct contained in the urine in a simple and quick manner. With this kit the ACR-lys adduct contained in the urine is determined as the amount of formyldehydro- piperidino -lysine (FDP-Lys).
Specifications
Assay principle: |
Competitive ELISA (detection: 450 nm) |
Specifity: |
Specific to ACR-Lysine adducts. |
Measuring range: |
1.56 - 200 nmol/mL |
Format: |
96 wells (18 samples in triplicate assays) |
Applications: |
Urine from human and animals |
Storage: |
Store at 4 - 10°C |
Expiry: |
12 months from the manufacturing date |
Required but not provided: |
Micropipet and chip (100 micro L, 1000 micro L). |
| Content of this kit |
|
| Fixed antigen plate: | 1 plate (96 wells) |
| Sample diluent: | 1 vial |
| Standards (8 levels) | 1 vial each. |
| First-order reaction antibody: | 1 vial |
| Diluent for first-order reaction antibody: | 1 vial |
| Stock solution for washing: | 1 vial |
| Enzyme-labeled antibody: | 1 vial |
| Diluent for enzyme-labeled antibody: | 1 vial |
| Coloring solution A: | 1 vial |
| Coloring solution B: | 1 vial |
| Short stop: | 1 vial |
| Plate seal: | 2 sheets |
| Assay Procedure | |
| 1) | Prepare samples and the standard solutions, fill the Antigen fixed plate with 50 micro L/well each. After the completion of sample filling, sway the whole plate gently in a manner so that the liquid comes into contact with the well uniformly. |
| 2) | Fill with 50 micro L/well of the previously prepared primary reaction antibody solution. After the completion of filling, sway the whole plate gently in a manner so that the liquid comes into contact with the well uniformly. Be sure to use a continuous filling instrument or a multichannel pipet. |
| 3) | Apply a plate seal and then allow to stand at room temperature (15 to 23°C) for 30 minutes. |
| 4) | Wash the plate with washings (4 times x 200 to 300 micro L/well). Finally, with light taps of a paper towel with the plate’s well side down to eliminate any water remaining in the well. |
| 5) | Fill with 100 micro L/well of the previously prepared secondary reaction antibody solution. After the completion of filling, sway the w hole plate gently in a manner so that the liquid comes into contact with the well uniformly. |
| 6) | Apply a plate seal and then allow to stand at room temperature (15 to 23°C) for one hour. |
| 7) | Wash the plate with washings (4 times x 200 to 300 micro L/well). Finally, with light taps of a paper tow el with the plate's w ell side down to eliminate any water remaining in the well. |
| 8) | Fill with 100 micro L/well of the previously prepared coloring solution. After the completion of filling, sway the w hole plate gently in a manner so that the liquid comes into contact with the well uniformly. |
| 9) | Apply a plate seal and then allow to stand at room temperature (15 to 23°C) for 5 to 15 minutes. Be sure to bring reaction to a halt so that the maximum absorbance reaches about 1.0 to 2.0 at that point in time. |
| 10) | After filling with 50 micro L/well of the short stop, sway the whole plate gently in a manner so that the liquid comes into contact with the well uniformly. Determine absorbance within 15 minutes of the reaction halt. |
| 11) | Determine absorbance at 450 nm using a plate reader. You may also be able to determine absorbance using a complementary wavelength of 590 to 620 nm. |
| 12) | Draw a calibration curve from the absorbance of the standard solution,
and calculate the ACR adduct protein amount, based on the absorbance of the sample. |
REFERENCES
-
Acrolein is a product of lipid peroxidation?formation of freeacrolein and its conjugate with lysine residues in oxidative low density lipoproteins.
Koji Utida et al., J.Biol.Chem.,273,16058(1998) -
Protein-bound acrolein ?Potential markers for oxidative stress.
Koji Utida et al., Proc.Natl.Acad.Sci.USA,95, 4882(1998) -
Protein-bound acrolein ?A novel marker of oxidative stress in Alzheimer's disease.
Noel Y.Calingasar et al., J.Neurochemistry,72, 751(1999) -
A 1-hour enzyme-linked immunosorbent assay for quantitation of acrolein and hydroxynonenal-modified proteins by epitope-bound casein matrix method.
K.Satoh, et al., Analytical Biochemistry,270, 323-328 (1999) -
Polyamine oxidase and acrolein as novel biochemical markers for diagnosis of cerebral stroke.
Tomitori M. et al., Stroke, 36, 2609 (2005)
The ‘Acrolein ELISA Kit’ is manufactured in Japan by the Life Science Division of the Nippon Oils and Fats Corporation, Japan. Genox is the distributor of this kit for users in North and South America.
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for RESEARCH USE ONLY.
Users of Genox’s
products are strongly advised NOT TO USE THEM FOR CLINICAL/DIAGNOSTIC
APPLICATIONS.
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