The Quantitative Determination of Vitamin B-12 Concentration in Human Serum by a Microplate Enzyme Immunoassay, Colorimetric.
Contents of Kit
1. Vitamin B-12 Calibrators – 1 mL/vial - Icons A-F
2. Vitamin B-12 Enzyme Reagent – 7.0 mL/vial – Icon
3. Vitamin B-12 Biotin Reagent – 7.0 mL/vial - Icon
4. Streptavidin Coated Plate – 96 wells – Icon
5. Wash Solution Concentrate – 20 mL/vial - Icon
6. Substrate Reagent – 12 mL/vial - Icon
7. Stop Solution – 8 mL/vial - Icon
8. Releasing Agent – 12 mL/vial – Icon
9. Stabilizing Agent – 0.5 mL/vial – Icon
10. Neutralizing Buffer – 7 mL/vial – Icon
Store the kit at 4°C upon receipt. For more detailed information, please download the following document on our website.
1. It is important that the time of reaction in each well is held constant to achieve reproducible results.
2. Pipetting of samples should not extend beyond ten (10) minutes to avoid assay drift.
3. Highly lipemic, hemolyzed or grossly contaminated specimen(s) should not be used.
4. If more than one (1) plate is used, it is recommended to repeat the dose response curve.
5. The addition of substrate solution initiates a kinetic reaction, which is terminated by the addition of the stop solution. Therefore, the substrate and stop solution should be added in the same sequence to eliminate any time-deviation during reaction.
6. Plate readers measure vertically. Do not touch the bottom of the wells.
7. Failure to remove adhering solution adequately in the aspiration or decantation wash step(s) may result in poor replication and spurious results.
8. Use components from the same lot. No intermixing of reagents from different batches.
9. Accurate and precise pipetting, as well as following the exact time and temperature requirements prescribed, are essential. Any deviation from Monobind's IFU may yield inaccurate results.
10. All applicable national standards, regulations and laws, including, but not limited to, good laboratory procedures, must be strictly followed to ensure compliance and proper device usage.
11. It is important to calibrate all the equipment e.g. Pipettes, Readers, Washers and/or the automated instruments used with this device, and to perform routine preventative maintenance.
The within and between assay precision of the Vitamin B-12 ELISA Test System were determined by analyses on three different levels of pool control sera. The number, mean values, standard deviation and coefficient of variation for each of these control sera are presented in Table 2 and Table 3.
*As measured in ten experiments in duplicate over a ten day period.
Vitamin B-12 is one of the nine water soluble vitamins important for healthy body functioning. The most important roles Vitamin B-12 plays in the human body are in the formation of red blood cells and the formation of the myelin sheath around the nerves. Since the effects are seen in body systems with a large range of function, the symptoms of Vitamin B-12 deficiency can sometimes be very ambiguous. A deficiency may also take from months to years to manifest depending on the cause and severity. 1, 2, 3 Two of the most common causes of Vitamin B-12 deficiency are diet and age. Because most sources of dietary Vitamin B-12 come from animals, vegans who do not efficiently supplement their diet are at risk. The elderly community is also at high risk because of their diet, as well as the less efficient functioning of their digestive system. 1, 3, 4 Intake of Vitamin B-12 starts by ingestion and then digestion by saliva. Once reaching the gut, Vitamin B-12 bound to proteins in food are released by the acids present. The B12 can then bind the Intrinsic factor. Once bound to IF, Vitamin B-12 is stable enough to travel into the intestines where it can be absorbed into your body through of its association with IF. 1, 5, 6, 7 Two very useful tests to distinguish between Vitamin B-12 deficiency and folate deficiency are methylmalonyl CoA (MMA) and homocysteine (hcy). Both deficiencies are represented by similar symptoms; however, even though both show increased levels of homocysteine, only Vitamin B-12 deficiency causes an increase in methylmalonyl CoA. The increase in levels of methylmalonyl CoA and homocysteine is thought to be the root cause of any symptoms that accompany a Vitamin B-12 deficiency. High levels of these two analytes in the blood stream causes increased oxidative stress to cells therefore causing increased apoptosis. In turn, vascular disease results in the form of atherosclerosis, coronary heart disease and/or neurodegeneration.