| ICMR approved AXIBIO - X-Spin Viral RNA ExtractionKit |
|---|
Components of the X-Spin Viral RNA Extraction Kit (50 Preps)
Cat No. XRS001050
| Sr.No. | Components | Quantity |
|---|---|---|
| 1 | VNE Buffer | 5 ml |
| 2 | Wash Buffer 1 * (concentrate) W1 | 11 ml (add ethanol 14 ml before use) |
| 3 | Wash Buffer 2 * (concentrate) W2 | 8 ml (add ethanol 17 ml before use) |
| 4 | Elution Buffer | 5 ml |
| 5 | Carrier RNA | 0.4 mg (Make final vol. 0.8 ml by adding elution buffer) |
| 6 | Spin Column | 50 Nos. |
| 7 | Collection Tube | 50 Nos. |
| 8 | User Manual | 1 |
* Compatible for Manual & Automated Machine Extractions.
Introduction and Intended Use
X-Spin Viral RNA Extraction Kit provides a fast an easy isolation method for the extraction and purification of nucleic acid (RNA) in samples. Its processed products are for clinical in-vitro diagnostics. The method uses a silica membrane column which along with buffer system allows efficient lysis followed by selective binding of RNA to the spin column and elusion of purified viral RNAusing centrifugation.
| Specification | |
|---|---|
| Principle | Mini spin column (silica matrix) |
| Sample | 200 µl cell-free fluid such as serum, plasma, body fluid and cell cultured supernatant or VTM. |
| Fragment Size | 100 bp ~30 kb |
| Recovery Rate | 80 ~ 90% |
| Binding Capacity | 30 µg |
| Elution Volume | 40 ~ 50 µl |
| Important Notes: | |
|---|---|
| 1 | Make sure everything is RNase-free when handling RNA. |
| 2 | Buffers provided in this system contain irritants. Wear gloves and lab coat when handling these buffers. |
| 3 | Add required ethanol (96-100% to Wash Buffer-1 and Wash Buffer-2 before use. |
| 4 | Preheat elution buffer to 56°C for elution step. |
| 5 | Add 4 ml Elution Buffer to Carrier RNAtube as per instruction. |
| General Protocol : | |
|---|---|
| 1 | Prepare pre-mixture of VNE buffer and carrier RNA by adding 16 µl carrier RNA per 100 µl VNE buffer. Alternatively, prepare the volume required based on the number of extractions required. Ex: For 5 RNA isolation, add 80 µl carrier RNA into 500 µl VNE buffer. |
| 2 | Add 100 µl of VNE buffer into micro-centrifugetube. |
| 3 | Add 200 µl VTM/sample to the VNE buffer in the micro-centrifuge tube. Mix by vortexing for 15-30 seconds. Note: To ensure efficient VNE, it is essential that the sample is mixed thoroughly with VNE buffer to yield a homogeneous solution. Frozen samples that have only been thawed once can also be used. |
| 4 | Incubate at room temperature (15-25°C) for 10 minutes. Note: Viral particle VNE is complete after VNE for 10 minutes at room temperature. No need for longer incubation times as it does not improve the yield or quality of the purified RNA. |
| 5 | Add 270 µl ethanol (96-100%) to the sample, and mix by vortexing for 15 seconds. After mixing, briefly centrifuge the tube to remove drops from inside the lid. |
| 6 | Carefully apply the entire lysed sample solution from Step 5 to the spin column (in a 2-ml collection tube). Close the cap, centrifuge at 8,000 rpm for 1 minute. Discard flow-through. |
| 7 | Add 500 µl Wash Buffer-1 and centrifuge at 8,000 rpm for 1 minute. Discard the flow-through. |
| 8 | Add 500 µl Wash Buffer-2 and centrifuge at 12,000 rpm for 1 minute. Discard the flow-through. |
| 9 | Remove the spin column from the tube and place it in a centrifuge tube and centrifuge at 12,000 rpm for 1 minute as a dry wash to remove the residual ethanol from the spin column. |
| 10 | Place the spin column in a clean 1.5 ml micro-centrifuge tube and add 50 µl Elution Buffer equilibrated to room temperature. Incubate at room temperature for 1 minute. Note: Use pre-warmed elution buffer (at 56°C). |
| 11 | The eluted RNA should be store at -80°C or below or use it immediately. |
Storage and Validity
AxiBio X-Spin Viral RNA Extraction Kit can be stored at room temperature (15-25°C) upto 12 months from the date of manufacturing. Please use before the expiry date mentioned on the product label.
Introduction and Intended Use
| Sr.No. | Problems | Possible reasons | Solutions |
|---|---|---|---|
| 1 | Low or no yield of genomic DNA Incorrect preparation of Wash Buffer 1 or Wash Buffer 2 | Wash Buffer W1 and Wash Buffer W2 is not mixed with ethanol before use | Make sure that the correct volumes of ethanol (96- 100 %) is added into Wash Buffer W1 and Wash Buffer W2 when first open. Repeat the extraction procedure with a new sample |
| The volume or the percentage of ethanol is not correct before adding into Wash Buffer W1 and Wash Buffer W2 | Make sure that the correct volumes of ethanol (96- 100 %) is added into Wash Buffer W1 and Wash Buffer W2 when first use. Repeat the extraction procedure with a new sample | ||
| 2 | Low or no yield of genomic DNA Elution of genomic RNA is not efficient | RNase-free water not completely absorbed by column membrane | After RNase-free water is added, stand the Spin Column for 2 min before centrifugation |
| 3 | Column is clogged | Sample is too viscous | Reduce the sample volume |
| 4 | Degradation of elutated RNA | Sample is old | Always use fresh or well-stored sample viral nucleic acid extraction |
