Description

The oasig freeze drying process stabilises all of the active components allowing these reagents to be shipped and stored at room temperature. This hugely simplifies the logistics of purchasing, shipping and using the technology. Whether you are in a sophisticated laboratory in Texas or a mobile field hospital in Timbuktu we can supply complete qPCR kit and reagent packages to your door quickly and cheaply via standard shipping methods without the need for dry ice or a cold chain of any sort.

The performance of the reagents is second to none. We are confident that you will find excellent data quality and even see an improvement in data quality versus many traditional frozen master mixes.

Primerdesign real-time PCR reagents are manufactured to the highest standards within our ISO9001:2008 and ISO13485:2012 certified quality management laboratory environment.

150 reaction pack of freeze dried oasig Lyophilised OneStep RT-qPCR Mastermix. 3 x 50 reaction vials and resuspension buffer.

Overview

• Detection kits for the BKV/JCV
• Available in TaqMan format for analysis

The BK virus is a member of the polyomavirus family. It has been suggested that this virus may be transmitted through respiratory fluids or urine, since infected individuals periodically excrete virus in the urine. BK viral infections are typically asymptomatic in healthy individuals, however very mild symptoms may appear including mild respiratory infections and fever. Infections with BK virus in immunocompromised or immunosupressed patients are much more severe and may involve renal dysfunction. In fact, in kidney transplant patients the immunosupressive drugs required for the transplant may allow the virus to replicate within the graft, resulting in a disease called BK virus nephropathy (BKVN). The JC virus is a type of human polyomavirus and is very common in the general population, infecting 70 to 90% of humans. Most people acquire JCV in childhood or adolescence. Typically the infection is subclinical and no of consequence in individuals with healthy immune systems. The initial site of infection may be the tonsils or the gastrointestinal tract, and the virus then remains latent in the gastrointestinal tract. JCV can also infect the tubular epithelial cells in the kidneys, where it continues to reproduce, shedding virus particles in the urine. Also, JCV can cross the blood-brain barrier into the central nervous system. JCV is known to cause the usually fatal progressive multifocal leukoencephalopathy (PML) by destroying oligodendrocytes in the brain in immunodeficient or immunosuppressed individuals. The JC and BK viruses are very similar, with their genomes sharing 75% homology. It is however important to differentiate between the viruses due to the differences in pathology and especially the invariably fatal outcome of PML which is only caused by the JC virus.

Details

Supporting Data

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Storage Conditions and Product Stability
All kit components can be stored for 2 years after the date of production without showing any reduction in performance.

All kit components should be stored at -20°C upon arrival.

Component	Cat. TM39350 (100 preps)	Cat. TM39310 (100 preps)
MDx TaqMan 2X PCR Master Mix	2 x 700 μL	–
BKV/JCV Primer & Probe Mix	280 μL	280 μL
BKV/JCV Positive Control	150 μL	150 μL
Nuclease-Free Water (Negative Control)	1.25 mL	1.25 mL
Product Insert	1	1

Introduction

Hipure viral RNA kit is suitable for purifying viral RNA from samples such as cell-free body fluid or culture medium. The kit is based on silica gel column purification technology. It requires no toxic phenol chloroform extraction and time-consuming alcohol precipitation in the extraction. The whole extraction process takes only 25 minutes. The kit is suitable for extracting viral RNA from 1-140µl cell-free fluid samples such as serum, plasma, cell-free body fluid or culture medium. The product has successfully extracted hepatitis B A/C, hepatitis C RNA, SARS and HIV. The obtained RNA can be directly used for RT-PCR, Northern hybridization and virus detection.

Details

Specifications

Features	Specifications
Main Functions	Extract viral RNA from 140μl cell-free samples
Applications	RT-PCR, Northern hybridization, and various virus detection
Purification method	Mini spin column
Purification technology	Silica technology
Process method	Manual (centrifugation or vacuum)
Sample type	Cell-free body fluid or culture medium
Sample amount	140μl
Elution volume	≥15μ
Time per run	≤25 minutes
Liquid carrying volume per column	800μl
Binding yield of column	100μg

Principle

Hipure silica gel column is based on glass fiber filter membrane with high binding force. Under the condition of high concentration of ionizing agent (such as guanidine hydrochloride or guanidine isothiocyanate), the filter membrane can adsorb nucleic acid through hydrogen bond and electrostatic, while protein and other impurities are not adsorbed and removed. The filter membrane adsorbed with nucleic acid is washed to remove the residual protein and salt. Finally, the nucleic acid adsorbed on the filter membrane can be washed out with low salt buffer (such as buffer TE) or water. The obtained nucleic acid has high purity and can be directly used in various downstream experiments.

The sample is homogenized and lysed in the lysate, and RNA is released into the lysate. The high concentration of guanidine isothiocyanate contained in the lysate denatured and inactivated endogenous or exogenous RNase, while RNA is protected from degradation. The lysate is centrifuged to remove insoluble impurities. After adding ethanol to adjust the binding conditions, it is transferred to the column for filtration. RNA is adsorbed on the membrane of the column, while protein is removed without adsorption. The column is washed with buffer VHB to remove protein and other impurities, washed with buffer RW2 to remove salt. Finally the RNA is eluted by RNase free water. The eluted RNA can be directly used in RT-PCR, Northern blot, poly-A purification, in vitro translation, etc.

Advantages

• Fast – several samples can be extracted in 20 minutes by column method
• High quality – high purity total RNA can be directly used in various sensitive downstream applications
• Safe – no phenol chloroform extraction required
• Sensitive – RNA can be recovered at the level of PG
• High yield – carrier RNA contained in the product maximize the recovery of trace nucleic acid

Kit Contents

Contents	R417102	R417103
Purification Times	50 Preps	250 Preps
HiPure Viral Micro Columns	50	250
2ml Collection Tubes	100	500
Buffer VRL	50 ml	200 ml
Carrier RNA	310 µg	3 x 310 µg
Buffer VHB*	13 ml	110 ml
Buffer RW2*	20 ml	2 x 50 ml
Nuclease Free Water	10 ml	30 ml

Storage and Stability

Carrier RNA should be stored at 2–8°C upon arrival. However, short-term storage (up to 12 weeks) at room temperature (15–25°C) does not affect their performance. The remaining kit components can be stored at room temperature (15–25°C) and are stable for at least 18 months under theseconditions

Hipure viral RNA kit is suitable for purifying viral RNA from samples such as cell-free body fluid or culture medium. The kit is based on silica gel column purification technology. It requires no toxic phenol chloroform extraction and time-consuming alcohol precipitation in the extraction. The whole extraction process takes only 25 minutes. The kit is suitable for extracting viral RNA from 1-140µl cell-free fluid samples such as serum, plasma, cell-free body fluid or culture medium. The product has successfully extracted hepatitis B A/C, hepatitis C RNA, SARS and HIV. The obtained RNA can be directly used for RT-PCR, Northern hybridization and virus detection.