Clostridium difficile toxin

A/B Assay for human feces Specifications Sheet

Background

C. difficile is a gram positive spore-forming bacteria and is the leading cause of hospital-acquired enteritis. In particular, pathogenic C. difficile is a causative agent of several antibiotic-associated diarrheal diseases commonly induced by treatment with antibiotics or by disruption of endogenous gastrointestinal flora. In the US alone, greater than 250,000 disease cases can be seen every year with associated annual healthcare costs exceeding US $1 billion.1 The worldwide burden is further troublesome. The main method of virulence and pathogenicity is due to expression of exotoxins, specifically by genes TcdA and TcdB. Both toxins, A and B specifically, are proinflammatory and cytotoxic resulting in fluid accumulation and extensive damage to the large intestine. 2 Improved detection of pathogenic C. difficile is recommended via PCR due to sensitivity and specificity. Fluxergy’s rapid diagnostic test is pathogen toxin specific and provides reliable detection in a point-of-use format.

References

  1. Carter GP, Rood JI, Lyras D. The role of toxin A and toxin B in Clostridium difficile-associated disease: Past and present perspectives. Gut Microbes. 2010;1(1):58-64.
  2. Voth DE, Ballard JD. Clostridium difficile Toxins: Mechanism of Action and Role in Disease. Clinical Microbiology Reviews. 2005;18(2):247-263.

Key Benefits

ic_check

Simple workflow

ic_check

No sample preparation

ic_check

Automated data interpretation

ic_check

Detection of both C. difficile toxin A and B

ic_check

Sample to result in 30 minutes

Assay Specifications

Sample Type Fecal Swab
Species Human
Storage Condition Fluxergy Card – Room temperature @ ~23° C
Fluxergy Buffers – Frozen @ 4° C

Kit Contents

  • 6” Rayon swab
  • Fluxergy C. difficile A/B Buffer 1
  • Fluxergy C. difficile A/B Buffer 2
  • Fluxergy Card

Customer supplied reagents and supplies

  • Clinical sample for testing

Sample collection method

Fecal swab. It is recommended that the head of the swab is fully coated with sample material. For optimal performance, collect feces to be tested in a sterile 2 oz specimen cup. Fully submerge the head of the swab into the sample for 5 seconds. Place stool coated swab into the swab tube. The opposite end of the tube serves as a tight-fitting cap. Place sample in fridge if planning to store for longer than 12 hours prior to testing.

Intended use

The Fluxergy clostridium difficile A/B assay is a qualitative, in vitro diagnostic for the rapid detection of pathogenic clostridium difficile toxins A and B in human fecal samples. This assay is for Research Use Only (RUO).

Positive PCR Result: indicates that DNA or RNA of the target organism(s) is present in the tested sample.

Negative PCR Result: indicates that DNA or RNA of the target organism(s) is not detected in the tested sample. However, a negative PCR results may also indicate that the number of target organisms is below the limit of detection.

Warnings and Precautions

  • Fluxergy’s clostridium difficile A/B assay is for Research Use Only (RUO). It is not intended for diagnostic use.
  • Fluxergy’s clostridium difficile A/B assay is compatible only with the Fluxergy Analyzer device.
  • All specimens should be handled as potentially infectious agents and according to universal safety precautions.
  • This Fluxergy clostridium difficile A/B assay is compatible only for human fecal swabs.
  • Contamination of the sample and kit contents may lead to erroneous results. Use aseptic technique and a clean workspace whenever possible.
  • Store Fluxergy assay kit at recommended storage temperature and conduct assay within specified environment (e.g. temperature and humidity) for optimal performance.
  • Follow appropriate specimen collection, storage and processing for optimal performance.
  • Use Fluxergy supplied swabs, reagents, pipettes and pipette tips to conduct assay for optimal performance.

Detection of Clinical Sample

Example amplification curve for clinical sample detection. PCR was conducted using the Fluxergy C. difficile A/B Assay C. difficile Toxin A and B specific primers and probes on a clinical sample acquired from iSpecimen. As a positive control, synthetic template at a concentration of 104 cp/mL was spiked into C. difficile Toxin A and B negative feces. Negative sample showed no amplification. Positive pathogen specific detection can be seen for both clinical and spiked samples.

Clinical Performance

Assay Performance

Lower Limit of Detection 103 cp/mL of either C. difficile toxin A or B
Sensitivity 100%
Specificity 100%

Positive and Negative Samples Tested. 100% agreement was shown between standard of care and Fluxergy results. Negative samples were acquired from Bioreclamation IVT.

Positive Negative
Standard of Care PCR C. difficile reflex test 10 10
Fluxergy Analyzer C. difficile toxin A/B assay 10 10

Table of Positive Samples Tested. 10 C. difficile Toxin A and B positive fecal samples were acquired from iSpecimen of varying age and sex. The Fluxergy Analyzer result agreed with the Standard of Care (SOC) result, specifically a C. difficile PCR reflex test, for each of the samples tested.

Sample ID Sample Type Analyte SOC Result Age Sex Fluxergy Analyzer Result
171218AAC0001 Stool Clostridium difficile toxin A+B Positive 63 F Positive
171218AAC0002 Stool Clostridium difficile toxin A+B Positive 70 M Positive
171218AAC0003 Stool Clostridium difficile toxin A+B Positive 88 M Positive
171218AAC0004 Stool Clostridium difficile toxin A+B Positive 77 M Positive
171218AAC0005 Stool Clostridium difficile toxin A+B Positive 59 F Positive
171218AAC0006 Stool Clostridium difficile toxin A+B Positive 28 F Positive
171218AAC0007 Stool Clostridium difficile toxin A+B Positive 50 M Positive
171218AAC0008 Stool Clostridium difficile toxin A+B Positive 72 M Positive
171218AAC0009 Stool Clostridium difficile toxin A+B Positive 67 F Positive
171218AAC0010 Stool Clostridium difficile toxin A+B Positive 68 M Positive