CoproELISA C. difficile GDH Assay Kit

$350.00

The Eagle Biosciences CoproELISA C. difficile GDH is an Enzyme-Linked Immunosorbent Assay (ELISA) for use as a screening test to detect Clostridium difficile glutamate dehydrogenase (GDH) in human fecal specimens from patients suspected of having C. difficile disease. The test does not distinguish between toxinogenic and nontoxinogenic C. difficile strains and is intended to aid in the diagnosis of C. difficile. Additional tests should be conducted to detect C. difficile toxins. The Eagle Biosciences CoproELISA C. difficile GDH Assay Kit is for Research Use Only and is not intended for diagnostic or therapeutic purposes.

CoproELISA C. difficile GDH Assay Kit

For Research Use Only

Size: 1×96 wells
Sensitivity: Cut-Off Control
Incubation Time: < 2 hours
Sample Type: Stool
Sample Size: 0.1 to 0.3g


Additional Information

Assay Background

The gram-positive anaerobic bacillus Clostridium difficile is the leading causative agent of antibiotic-associated diarrhea and pseudomembranous colitiss. This pathogen is capable of causing disease that could be severe or fatal if not diagnosed on time and treated. Exposure to antibiotics is the major risk factor for C. difficile infection. Infection can develop if the normal gastrointestinal flora is disrupted by antibiotic therapy and a person acquires toxin-producing C. difficile, typically via the fecal-oral route. C. difficile’s key virulence factors are toxin A and toxin B. These toxins show high sequence and functional homology. Toxin A has been described as a tissue damaging enterotoxin which attracts neutrophils and monocytes and toxin B as a potent cytotoxin that degrades the colonic epithelial cells. Most virulent strains produce both toxins, however, toxin A negative/toxin B positive strains are also capable of causing disease. All strains of C. difficile produce high levels of GDH. Therefore, C. difficile’s GDH enzyme is considered a very good antigen marker for detection of this organism. The CoproELISA C. difficile GDH test is a highly specific and sensitive diagnostic kit for GDH in stool specimens. A positive result confirms the presence of C. difficile and a negative result indicates its absence. A separate test should be performed to confirm the presence of C. difficile toxins in the GDH positive samples.

Assay Principle

Break-apart microwells are coated with C. difficile GDH specific monoclonal antibodies. A horseradish peroxidase (HRP) conjugated monoclonal anti-GDH antibody is added to the antibody-coated microwells. Fecal samples are diluted in sample diluent and added to the microwells. In this step C. difficile GDH is marked by the HRP conjugated antibody and immobilized by the coated antibody. Unbound conjugate is removed by washing. TMB-substrate is added, the substrate is hydrolyzed by the peroxidase and yields a blue solution of the reduced substrate. Upon the addition of the stop solution, the blue color turns yellow and should be read by an ELISA reader at a wavelength of 450/620 nm. The absorbance is proportional to the level of C. difficile GDH in the sample.

Cross Reaction

The CoproELISA C. difficile GDH test was evaluated using microbial culture isolates and clinical stool specimens*. No cross-reactivity was observed with any of the gastrointestinal pathogens and microbes listed: Blastocystis, Campylobacter, Cryptosporidium parvum, Dientamoeba fragilis, Escherichia coli, Entamoeba histolitica, Enterococcus faecali, Enterococcus faesium, Enterococcus avium, Enterococcus aerogenes, Enterococcus cloacae, Enterococcus gallinarum, Enterococcus durans, Giardia lamblia, Helicobacter pylori, Klebsiella pneumonia,
Salmonella enterica, and Shigella.

References

  1. Cloud J, Kelly CP. Update on Clostridium difficile associated disease. Curr Opin Gastroenterol. 2007 Jan;23(1):4-9.
  2. Owens RC Jr, Donskey CJ, Gaynes RP, Loo VG. Muto CA. Antimicrobial-associated risk factors for Clostridium difficile infection.Clin Infect Dis. 2008 Jan 15;46 Suppl 1:S19-31.
  3. Kelly CP, Pothoulakis C, LaMont JT. Clostridium difficile colitis. N Engl J Med. 1994 Jan 27;330(4):257-62
  4. Voth DE, Ballard JD. Clostridium difficile toxins: mechanism of action and role in disease. Clin Microbiol Rev. 2005 Apr;18(2):247-63.
  5. Savidge TC, Pan WH, Newman P, O’brien M, Anton PM, Pothoulakis C. Clostridium difficile toxin B is an inflammatory enterotoxin in human intestine. Gastroenterology. 2003 Aug;125(2):413-20.
  6. Pituch H, van den Braak N, van Leeuwen W, van Belkum A, Martirosian G, Obuch-Woszczatyński P, Łuczak M, Meisel-Mikołajczyk F. Clonal dissemination of a toxin-A-negative/toxin-B-positive Clostridium difficile strain from patients with antibiotic-associated diarrhea in Poland. Clin Microbiol Infect. 2001 Aug;7(8):442-6.
  7. Shin BM, Kuak EY, Yoo SJ, Shin WC, Yoo HM., Emerging toxin A-B+ variant strain of Clostridium difficile responsible for pseudomembranous colitis at a tertiary care hospital in Korea. Diagn Microbiol Infect Dis. 2008 Apr;60(4):333-7.
  8. Lyerly DM, Barroso LA, Wilkins TD. Identification of the latex test-reactive protein of Clostridium difficile as glutamate dehydrogenase. J Clin Microbiol. 1991 Nov;29(11):2639-42.
  9. Carman RJ, Wickham KN, Chen L, Lawrence AM, Boone JH, Wilkins TD, Kerkering TM, Lyerly DM. Glutamate dehydrogenase is highly conserved among Clostridium difficile ribotypes. J Clin Microbiol. 2012 Apr;50(4):1425-6.

Manual

Product Manual