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Hc2 GC-ID DNA Test
An in vitro nucleic Acid Hybridization Microplate
Assay with Amplification using Microplate chemiluminescene for the
Qualitative Detection of Neisseria Gonorrhoeae (GC) DNA in
cervical Specimen’s.
Summary and explanation
Neisseria gonorrhoeae are non-motile. Gram negative
diplococci with fairly complex growth requirements. They are
aerobic, producing optimal growth at temperatures in the range of
35-37 °c in the presence of 37% co2 and ≥70% relative humidity.
Presumptive diagnosis for Neisseria gonorrhoeae is traditionally
obtained by isolating organisms from cultures of clinical
specimens and using a gram stain for morphological examination.
Definitive diagnoses can be obtained with a positive oxidase
and/or catalase test of the culture. Additional confirmation of
results includes carbohydrate degradation, agglutination, and
sugar fermentation tests. More definitive, direct test for
Neisseria gonorrhoeae include antigen detection and nucleic acid
probe tests. An enzyme linked immunosorbent assay has been shown
to be as sensitive and as specific as the gram stain for detecting
gonococci in male urethral and first void urine specimens, but it
has decreased sensitivity when applied to endocervical specimens.
Because the antigen detection test may cross-react with commensal
Neisseria and related species. This test can only be used for
presumptive diagnosis.
More recently, nucleic acid hybridization tests
have been used to evaluate clinical samples for the detection of
Neisseria gonorrhoeae in high-risk populations using both
endocervical and male urethral specimens.
PRINCIPLE OF THE PROCEDURE
The hc2 GC-ID DNA Test using Hybrid Capture 2
technology is a nucleic acid hybridization assay with signal
amplification that utilizes microplate chemiluminescent detection.
Specimens containing the target DNA hybridize with a specific GC
RNA probe. The resultant RNA:DNA hybrids are captured onto the
surface of a microplate well coated with antibodies specific for
RNA:DNA hybrids. Immobilized hybrids are then reacted with
alkaline phosphatase conjugated antibodies specific for RNA:DNA
hybrids, and detected with a chemiluminescent sunstrate. Several
alkaline phosphatase molecules are conjugated to each antibody.
Multiple conjugated antibodies bind to each captured hybrid
resulting in substantial signal amplification. As the substrate is
cleaved by the bound alkaline phosphatase, light is emitted, which
is measured as relative light units (RLUs) on a luminometer. The
intensity of the light emitted denotes the presence or absence of
target DNA in the specimen.
An RLU measurement equal to or greater than a
specified ratio to the positive cutoff (CO) value indicates the
presence of GC DNA in the specimen. An RLU measurement less than a
specified ratio to the positive cutoff value indicates the absence
of GC DNA or DNA or GC DNA levels below the detection limit of the
assay.
The GC probe contains a probe mixture specifically
chosen to eliminate or minimize cross-reactivity with DNA
sequences from human cells, other bacterial species, or Nesseria
SPECIES OTHER THAN Neisseria gonorrhoeae. The GC probe supplied
with the hc2 GC ID DNA Test is complementory to approximately.
High volume sample throughout testing with the hc2
GC ID DNA Test can be performed utilizing a general use automated
pipetting and dilution system referred to as the rapid capture
system (RCS). This instrument, using an application specific to
the hc2 GC-ID DNA test, processes up to 352 specimens in eight
hours. To enable high volume sample-throughput testing, all the
procedural steps of the assay are performed by the RCS, with the
exception of specimen denaturation, chemiluminescent signal
detection, and result reporting.
REAGENTS AND MATERIALS PROVIDED
There are 96 tests in one hc2 GC-ID DNA test kit .
the number of patient results will vary, depending on the number
of uses per kit.
1 use = 88 patient results
2 use = 80 patient results
3 use = 72 patient results
4 use = 64 patient results
1 * 0.35 ml Indicator Dye INDIC :
Contains 0.05% w/v sodium azide.
1 * 50 ml Denaturation Reagent REAG
DENAT: Dilute sodium hydroxide (NaOH) solution.
1 * 5 ml Probe Diluent DIL PROBE:
Buffered solution with 0.05% w/v sodium azide.
1 * 200 µl CT Probe PROBE CT: CT
RNA probe in buffered solution.
1 * 2 ml Negative Control CONTROL -
: Carrier DNA in specimen Transport Medium (STM) with 0.05% w/v
sodium azide.
1 * 1 ml CT Positive Calibrator (PC)
CAL CT + : 1.0 PG/ML cloned CT DNA and carrier DNA in STM
with 0.05% w/v sodium azide.
1 * 1 ml Quality Control GC (QC CT)
QC CT : 5.0 pg.ml cloned CT DNA and carrier DNA in STM
with 0.05% w/v sodium azide.
1 * 1 ml Quality Control GC (QC GC)
QC GC : 5.0pg/ml cloned GC DNA and carrier DNA in STM with
0.05% w/v sodium azide.
1 each Capture Microplate PLATE
CAPTURE : Coated with anti-RNA:DNA hybrid antibodies.
1 * 12 ml Detection Reagent 1 REAG DET
1 : Alkaline phosphatase-conjugated antibodies to RNA:DNA
hybrids in buffered solution with 0.05% w/v sodium azide.
1 * 12 ml Detection Reagent 2 REAG DET
2 : CDP-Star with Emerald II.
1 * 100 ml Wash Buffer Concentrate BUF
WASH X 30 : Contains 1.5% w/v sodium azide. |
