sina fungiplex universal-25T-PQ3681

Sina Fungiplex Universal
Real-Time PCR Kit

Kit component
Storage and Stability
Sina Fungiplex Universal is a ready to use kit and should be stored at -20 °C upon arrival.
Check Kit on arrival. If the packaging is damaged the kit must not be used. Additionally, all reagents contained
within the kit are shipped frozen and should arrive frozen.
The performance of the kit will be guaranteed until the printed expiration date, when the contents keep in an
appropriate condition.
Kit Description
Sina Fungiplex Universal real time PCR Kit is an in vitro diagnosis kit for the qualitative detection of Fungal DNA extracted
from respiratory tract specimens (BAL, BL, sputum) and body fluids (peritoneal, pleural and CSF) whole blood. The assay is
compatible with common laboratory real time PCR equipment result in less than 2 hours. This kit is ready to use reporting
format, are detailed in below:
Pophysomyces Cunninghamella Mucor Saccharomyces
Aspergillus Exophiala Paecilomyces Saksenaea
Candida Fusarium Penicillium Scedosporium
Cladosporium Histoplasma Pneumocystis Syncephalastrum
Coccidioides Lichtheimia Rhizomucor Talaromyces
Cryptococcus Microsporum Rhizopus Trichosporon
The rapid, and reliable detection provided by this kit is guaranteed by quality control processes that ensures the user can have
full confidence in assay quality and reproducibility.
WARNINGS AND PRECAUTIONS:
Chemical risks
There are no hazardous materials included in the manufacture of the Sina Fungiplex Universal real time PCR Kit. The
composition of all reagents represents no specific risk to the user or to their property.
Additional chemicals and materials may be required for procedures described in these Instructions for use. Carefully read any
warnings, instructions, or Safety Data Sheets provided by the supplier and follow general safety regulations when you hand
chemicals, biohazards, or other materials.
Description QNTY
(μl)
NO. Storage
(°C)
PCR Master Mix 625 1 -20
Primer Mix 375 1 -20
Positive control 50 1 -20
Biological risks
Sina Fungiplex Universal real time PCR Kit involves safe material, but working with Fungi is potentially
dangerous and transmissible. All personnel are responsible for reading and following all necessary health and
safety precautions.
It is very important to wear appropriate PPE at all times; a lab coat, protective gloves and safety glasses are
minimum needed for working with these materials.
Additional materials required (not provided)
General equipment
• Nucleic acid extraction kit SinaPure DNA (EX6011);
• Validated real-time thermal cycler instrument
• Class II hood / PCR station
• Vortex mixer
• Mini centrifuge
• Centrifuge capable of microplate processing
(recommended)
• Fridge (2 to 8°C) and freezers (-25°C to -15°C)
• Micropipettes for volumes of 1 to 1000 μL
Reagents
• Negative extraction control, for example nuclease free water (recommended);
Consumables
• Appropriate DNase / RNase free plastic-ware for
PCR preparation
• DNase / RNase free pipette tips
• Disposable gloves, powder-Free
• PCR plastic-ware compatible with the thermal
cycler of choice
Important notes: please read before starting
▪ Follow GLP rules when performing nucleic acid extraction from biological species.
▪ Check the procedure with a known Positive Control, if use the kit for the first time.
▪ It is important that the background signal should be considered while you analysis the samples and their
measurement should be excluded
Protocol:
Remove the Sina Fungiplex Universal real time PCR Kit from the freezer and allow to thaw. Quickly vortex the tubes once
defrosted. Prepare the final reaction mix as follows:
PCR Regent:
Reagent Volume
(µL)
Master Mix 2X 25
Primer/probe mix 15
Temp 10
Total 50
qPCR program:
Step Temperature (°C) Time Number of cycles Analysis mode Data collection
Pre-incubation 95 300 sec 1 None None
Denaturation 95 60 sec
40 Quantification
None
Annealing 61 25 sec Acquire
extension 72 20 sec

Analysis
Undet: Undetermined.

Tips and Suggestions
Positive and negative controls At least one positive and negative control should be included in each analysis. All control should
be treated and test in the same manner as patient samples. A negative control that yields a positive test result is indicative of a
mainly is due to cross contamination. The assay run should be repeated using a fresh aliquot of negative control material,
ensuring that the work area and equipment are properly decontaminated and extreme care is taken during PCR set-up. A false
negative result is indicative of reagent failure or sample handling error. Ensure all reagents have been stored correctly and,
where applicable, expiry date before repeating the assay run taking extreme care during PCR set-up. Expected results for the
positive control are provided.
Storage temperature
Sina Fungiplex Universal real time PCR Kit is shipped on dry ice and should be
stored within this advised temperature range immediately on receipt.
Troubleshooting
panfugi (Cyber)
≤ 40 Fungi detected.
Undet/blank or >40 Fungi not detected.
Undet/blank or >40 Invalid. Sample needs to be retested by repeating PCR if there is
sufficient extracted DNA.
Otherwise, sample needs to be retested by re-extraction or recollection from the original source.
Problem The assay is Insufficient amplification
Possible Causes
• The initial amount of template is too low.
• Thermal cycler was not at correct temperature
Diagnostic Test
• Check DNA absorbance ratio at 260 and 280 nm (260/280); Pure DNA should has a 260/280 ratio
• Check optimal conditions for your thermal cycler
Uniformity of Ct values across different tests
The clinical significance of positive results with high Ct are difficult to interpret in the absence of clinical history and
context. Positive results with low DNA load (high Ct) can be seen in the early stages of infection (before the person
becomes capable of infection transmission) or late occurrence of infection when the risk of transmission is low.
Kit Quality Control:
All components of this Kit are successfully tested in term of DNA purification and amplification reaction.