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Vitamin B12 ELISA kit blog

Vitamin B12 ELISA kit

At Los Angeles-based Diagnostic Automation (focusing on ELISA Kits) we offer two vitamin B12 elisa kits:

1) Food Vitamin B12 ELISA kit to determine vitamin B12 quantitatively in vitaminated food in a significantly faster way (2.5 to 4 hours inclusive sample pretreatment) compared to a conventional microbiological assay (24 to 48 hours)

2) Vitamin B12 ELISA kit to measure the serum concentration of vitamin B12

Vitamin B12, also known as cobalamin, has the largest and the most complicated structure in vitamins.  Cobalamin is an organometallic compound and only known essential molecule with a stable metal-carbon bond.  To discover vitamin B12, elucidate its metabolic roles, discover the signs and symptoms of its deficiency, and to find treatment, took over 100 years [1].  In 1850s Thomas Addison an English physician described a lethal (pernicious) anemia.  In 1930s Castel an American physician discovered the intrinsic factor (IF) essential for B12 absorbance and many years later it was found to be a glycoprotein.  In 1934 Whipple, Minot, and Murphy shared the Nobel Prize for their discovery of treating anemia with liver extract.  In 1950s Dorothy Hodgkin a British chemist described the structure of vitamin B12 and received the Nobel Prize for it in 1964[2].  Different biochemical roles of B12 such as its interaction with folate and their linked role in megaloblastic anemia, was identified subsequently.  B12 continues to be the subject of intense interest in research especially the deficiency in infants secondary to maternal origins and its role in their development [3].

 

 

Molecular structure of Vitamin B12 (Cobalamin)

Cobalamin/vitamin B12 is a water-soluble vitamin with a key role in the normal formation of red blood cells and also normal functioning of the nervous system including brain.  Moreover, it is involved in the DNA regulation; synthesis and fatty acid metabolism, thus effecting energy production in the body.  Every cell of human body needs vitamin B12 for its normal function.  Animal, Plants, and fungi are not capable of producing vitamin B12.  Only archaea and bacteria have the necessary enzymes required for the synthesis of B12 in the form of hydroxocobalamin which is then converted to cobalamin in the human body.  B12 is naturally present in animal foods and can also be received through supplements.  However, proper absorption in the body requires a complex pathway [4].

Vitamin B12 has profound effects on human health and is essential for several crucial functions in the body.  The deficiency manifest when serum levels are below the cutoff point.  In USA the limit is 200 picograms per milliliter (pg/ml) [5].  In Europe and Japan the limit for B12 is 500-550 pg/ml, this level of B12 may explain the lower rate of Alzheimer in these countries compared to USA with lower cut off (200pg/ml) [6].  It is essential to identify the B12 deficiency promptly.

Diagnostic Automation offers a rapid and reliable tool as an ELISA kit for measuring the serum concentration of vitamin B12.  Deficiency of vitamin B12 can manifest with hematological disorder, such as severe anemia requiring blood transfusions, and behavioral changes to serious and permanent nerve damage, neurologic, vascular, and gastrointestinal problems.

Vitamin B12 was discovered through investigation of Pernicious anemia.  Pernicious anemia is a member of the large family of megaloblastic anemias that is characterized by production of large immature and dysfunctional red blood cells in the body.  Pernicious anemia is a form of autoimmune disease that destroys cells in the stomach which are responsible for the production of Cubilin, an intrinsic factor (IF) which is a protein essential for the absorption of B12.  IF is located within the epithelium of intestine and kidney. Mutations in CUBN (Cubilin gene) may play a role in autosomal recessive megaloblastic anemia [7].  Real pernicious anemia constitutes only a small fraction of the cases, however, malabsorption due to diminished secretion of gastric acid and enzymes necessary for the cleavage of protein- B12 bond from food is the main concern.  It is very important to note that maintaining an optimal status of vitamin B12 is not only dependent on adequate dietary intake but more critically on its effective absorption which declines with age.

Vitamin B12 is absorbed through saliva at the ingestion stage, and then during digestion.  The vitamin B12 from food proteins is released with the help of acids in the stomach.  Absorption of B12 depends on intrinsic factor (IF) which is present in the gastric juice.  IF combines with vitamin B12 and facilitates its absorption in the lower part of the small intestine. Vitamin B12 and Folate deficiencies have similar symptoms, so it is important to have accurate tests that can detect the differences.  Fortunately, Vitamin B12 deficiency can easily be treated and prevented [8].  It is very important to be aware of it since there is a high prevalence of mild, subclinical B12 deficiency in asymptomatic individuals, especially with individuals at high risk such as vegan/vegetarian [9].

Delay in the diagnosis and treatment of vitamin B12 deficiencies can lead to development of severe and irreversible neurologic damage.  Although vitamin B12 deficiencies was estimated to occur at a frequency of 1 in every 31 adults 51 years of age or older in USA population, but the incidence of vitamin B12 deficiency is far more common than previously assessed [10; 11].  The rise in gastric bypass surgeries in obese women and growing incidence of gastrointestinal issues such as coeliac disease, Crohn’s, and ulcerative colitis, plus the increasing number of vegan/vegetarian mother are the growing concern [12].  These conditions substantially decrease the absorption of vitamin B12 which adversely affects the development of children born of these women. The association between vitamin B12 and cognitive development in children has been observed in infant born to vegan and vegetarian mothers.

A recent study in infants with B12 deficiency has reported diverse clinical abnormities including hypotonic muscle, apathy, cerebral atrophy, and nerve cell demyelination.  Some of these symptoms were reversible since, a rapid improvement in neurological symptoms was reported after vitamin B12 therapy.  However, in long term follow up many of these infants showed serious delay in cognitive and language development.  Babies born to vegetarian mothers may show deficiency of vitamin B12.  Delayed detection and treatment in infants with vitamin B12 deficiencies, can lead to severe and permanent damage to their nervous system.

At Diagnostic Automation (focusing on “ELISA Kits”) we offer vitamin B12 elisa kit which informs mothers before their pregnancy and upon the detection of low level of B12 using vitamin B12 elisa kit, supplement intake can prevent the complications due to deficiency if that is the primary cause of the deficiency.  Although our vitamin B12 serum elis kit is very fast and accurate, Homcysteine and methylmalonic acid can also be used to confirm a vitamin B12 deficiency for cases with ambiguous initial results because metabolic changes often precede low cobalamin levels [13].

Homocysteine levels in blood are influenced by blood levels of B-complex vitamins: cyanocobalamin (B12), folic acid (B9) and pyridoxine (B6).  Studies have shown that high plasma total homocysteine levels are associated with an increased risk of atherosclerotic diseases. Elevated level of homocysteine which is an amino acid with biological functions in methionine metabolism is a risk factor for cardiovascular events.  Cardiovascular disease, including coronary artery disease, stroke and congestive heart failure, is the number one cause of death worldwide.  Thus, it has been suggested that B vitamin supplementation might reduce the risk of myocardial infarction and stroke.

Preventive strategies might include healthy people with low or high risk for developing cardiovascular disease and people with an established cardiovascular disease to take these supplements.  A study including 12 randomized clinical trials involving 47,429 participants show no evidence that homocysteine lowering interventions such as supplementations of vitamins B12, B9 or B6, prevent myocardial infarction and stroke.  However, multiple studies have shown a significant improvement.

Heart Outcomes Prevention Evaluation 2 (HOPE 2) trial reanalysis, showed a reduced incidence of nonfatal stroke with long term (over 3 years) treatment with B vitamins [14-16].  Patients with homocystinuria are prone to thromboembolic events even with minor surgeries.  Levels of homocysteine should be reduced to a near normal level before any surgery.   Increased levels of methylmalonyl CoA and homocysteine in sever form can affect the vascular system and increase the diseases such as Parkinson, atherosclerosis, heart diseases, and stroke. It is postulated that the effect might be through increased level of stress at the cellular level, eventually leading to apoptosis (programed cell death).

Vitamin B12 has high level of importance in public health, not only because of its deficiency leading to megaloblastic anemia and irreversible nerve damage, but also because there are emerging evidence that links vitamin B12 deficiencies to an increased risk of a number of age related diseases, such as cardiovascular disease, cognitive dysfunction, and dementia.  Moreover, there are concerns relating the imbalance of B12 and folate, especially the potential adverse effects for older adults with low vitamin B12 status and over exposure to folate especially in countries with mandatory fortification of food with folate. Thus, inter relationship between vitamin B12 and folate is an important emerging issue in aging population.

Vitamin B12 deficiency can easily be treated and prevented.  There are inexpensive treatment options available as B12 supplement to treat a vitamin B12 deficiency. The treatment is safe, effective, with no known toxicity. To prevent B12 deficiency, all patients 51 years of age and older should be recommended to take B12 supplement daily however, the dosage recommendation may vary.  Recent studies show that in people with malabsorption problems of B12, high dose of oral or nasal intake may be as effective as injection.  However, in severe cases of deficiency such as pernicious anemia and neurological symptoms injection is still the preferred method of administrating vitamin B12.  In USA cyanocobalamin is the most frequent form of B12 supplementation, however recent research prefers hydroxycobalamins which is more common in Europe and it has shown to be superior to cyanocobalamin.

Finally, to identify B12 deficiency the first step is to take an accurate test to determine the level of B12 in the serum.  At CORTEZ DIAGNOSTICS we offer a rapid and reliable vitamin B12 deficiency elisa test kit proved to be both accurate and fast in determining B12 concentration in blood.  The next step is identifying the mechanism and problem leading to B12 deficiency.  This is a challenging step and requires professional help, after that the suitable form of B12 in a form of injection, oral, nasal, and sublingual and the dose and length of treatment will be determined easily.  All patients with unexplained hematologic or neurologic signs and symptoms should be evaluated for B12 deficiency since it is easy to miss the B12 deficiency.

 

Indications for B12 testing:

Megaloblastic or Pernicious anemia: a condition in which there is a lack of an intrinsic factor (IF)

Hyperhomocysteinaemia: an inherited disorder of the metabolism the amino acid methionine and Homocystinuria: also known as cystathionine beta synthase deficiency, both are associated with atherosclerosis, venous and arterial thrombosis.  Acquired forms can be treated with vitamins B12 and B6 and folate

Peripheral neuropathy: damage to the nerve of the peripheral nervous system, may be caused by diseases or trauma to the nerve or as a side effect of a systemic illness

Patients with GI problems: patients with gastritis (long term use of acid-reducing drugs), gastric surgery (e.g., gastric bypass, gastrectomy), gluten intolerance or intestinal resorption disorders, atrophic gastritis (a thinning of the stomach lining that affects up to 30% of people aged 50 and older)

Conditions affecting the small intestine: such as Crohn’s disease, Celiac diseases, parasite (B12 utilizing fish tapeworm such as the Diphyllobothrium latum) or bacterial overgrowth in the small intestine and viral infection (e.g. HIV)

Alcoholism: decrease vitamin B12 absorption

Autoimmune disorders: such as Grave’s disease or systemic lupus erythematous

Pancreatic insufficiency

Patients with thrombosis

Chronic liver and kidney disease

Vitamin B12 deficiency from diet: vegan and vegetarians; pregnancy and lactation

 

 

References:

1. Scott J.M. Molloy A.M. The Discovery of Vitamin B12. Ann Nutr Metab 2012;61:239–245

2. Dorothy Crowfoot, et al. Structure of Vitamin B12: The Crystal Structure of the Hexacarboxylic Acid derived from B12 and the Molecular Structure of the Vitamin. Nature 176, 325 – 328 (20 August 1955)

3. Pepper MR, Black MM . B12 in fetal development. Semin Cell Dev Biol. 2011, 22(6):619-23

4. Jeremy M Berg, John L Tymoczko, and Lubert Stryer.  Biochemistry, 5th edition W H Freeman; 2002. ISBN-10: 0-7167-3051-0

5. Goodman M, Chen XH, Darwish D. Are US lower normal B 12 limits too low? 1996. J Am Geriatr Soc 44 (10): 1274–5

6. Mitsuyama Y, Kogoh H. Serum and cerebrospinal fluid vitamin B12 levels in demented patients with CH3-B12 treatment–preliminary study. 1988. Jpn. J. Psychiatry Neurol.42 (1): 65–71

7. Stabler SP, Allen RH. Megoblastic anemias. In: Goldman L, Ausiello D, editors. Cecil Textbook of Medicine. 22nd ed. Philadelphia: W. B. Saunders Company; 2004. p. 1050-7

8. Medline plus, B12 deficiency: https://www.nlm.nih.gov/medlineplus/ency/article/000574.htm

9. de Benoist B.  Conclusions of a WHO Technical Consultation on folate and vitamin B12 deficiencies. Food Nutr Bull. 2008; 29(2 Suppl):S238-44

10. den Elzen WP, Gussekloo J. Anaemia in older persons. Front Hum Neurosci. 2013; 7: 97

11. Hughes CF, Ward M, Hoey L, McNulty H. Vitamin B12 and ageing: current issues and interaction with folate.  2013 April 16; Ann Clin Biochem.

12. Dror DK, Allen LH. Effect of vitamin B12 deficiency on neurodevelopment in infants: current knowledge and possible mechanisms. Nutr Rev. 2008; 66(5):250-5.

13. Sally P. Stabler. Vitamin B12 Deficiency. N Engl J Med. 2013; 368:149-160

14. Toole JF. et al. Lowering homocysteine in patients with ischemic stroke to prevent recurrent stroke, myocardial infarction, and death: the Vitamin Intervention for Stroke Prevention (VISP) randomized controlled trial. JAMA. 2004;291(5):565-75

15. Spence JD. Homocysteine-lowering therapy: a role in stroke prevention? Lancet Neurol. 2007;6(9):830-8

16. Saposnik G. et al. Homocysteine-lowering therapy and stroke risk, severity, and disability: additional findings from the HOPE 2 trial. Stroke. Apr 2009;40(4):1365-72