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Total RNA Purification Kit for Leukocytes, 96-well format Add to Cart

Cat#: 37800-NB
Quantity: 2 plates/192 preps
Price: 832 €
Supplier: Norgen
Shipping: RT
User Manual  

Rapid high throughput extraction and purification of total RNA and microRNA from leukocytes

• Isolate total leukocyte RNA - All leukocyte RNA species are isolated, from large mRNA down to microRNA.
• Fractionate leukocytes from whole blood in minutes - Rapid removal of red blood cells from whole blood samples using differential red blood cell lysis.
• Rapid and convenient 96-well plate format for high throughput applications
• No phenol or chloroform extractions

Norgen’s Leukocyte RNA Purification Kit provides a rapid method for the high throughput isolation and purification of total leukocyte RNA from mammalian blood samples. Selective isolation of leukocyte RNA results in improved expression profiling and other downstream applications by removing the masking effects of some RNAs which are very abundant in whole blood, such as globin mRNAs. The kit is able to isolate total leukocyte RNA, including both large mRNA and all small RNA species containing microRNA (miRNA) and small silencing RNA (siRNA). The purified RNA is of the highest quality and can be used in a number of downstream applications including real time PCR, reverse transcription PCR, northern blotting, RNase protection and primer extension, and expression array assays.

Purification is based on 96-well column chromatography using Norgen’s proprietary resin as the separation matrix. Purification can be performed using either a vacuum manifold or centrifugation. Briefly, the red blood cells are first removed from the sample through differential red blood cell lysis, and the leukocytes are recovered through centrifugation. The leukocytes are lysed, and the leukocyte RNA is bound to Norgen´s 96-well filter plates (BIND). Under these conditions only the RNA will bind to Norgen’s resin, while the DNA, proteins and other contaminants will be removed in the flowthrough. The bound RNA is then washed to remove any remaining impurities (WASH). Lastly, the purified RNA will be eluted into the provided Elution Buffer or water (ELUTE). Please see the procedure flowchart below.

Workflow
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Consistent Yield of High Quality Leukocyte RNA with Complete Size Range without the Use of Phenol. Norgen´s Leukocyte RNA Purification 96-Well Kit allows for the consistent isolation of high quality RNA, with complete size range from the very large RNA down to small RNA without the use of phenol. Total RNA was isolated from 200 µL aliquots of hamster blood using Norgen’s Leukocyte RNA Purification 96-Well Kit in 12 replicates. Five microliters of the 75 µL isolated RNA was resolved on a 1.2% formaldehyde agarose gel (Panel A) and 1 µL of the 75 µL isolated RNA was resolved on an Agilent RNA Nano 6000 Chip in a 2100 Bioanalyzer (Panel B). All 12 replicates showed both high yield and high quality. In addition, all replicates showed effective recovery of all sizes of RNA including the small RNA (arrow).


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Isolation of Both Large and Small RNA. Norgen’s Leukocyte RNA Purification 96-Well Kit allows for consistent isolation of both large and small RNA. Leukocyte RNA was isolated from 200 µL of hamster blood using Norgen’s Leukocyte RNA Purification 96-Well Kit. Aliquots of each total RNA sample were then used in 2 different RT-qPCR reactions. Ten microliters of the 75 µL isolated RNA was subjected to a 20 µL reverse transcription using miR-21 stem-loop reverse primer or oligo dT. Three microliters of the reverse transcription was used in a 20 µL real-time PCR reaction with primers to detect the miR-21 (Panel A) and the beta-actin transcripts (Panel B), respectively. Both the mRNA and microRNA were amplified in a consistent manner from all the samples, with low variability of the Ct values. Thus both types of RNA are being consistently isolated using this kit.


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