Leukocyte RNA Purification Kit

Supplier
Norgen Biotek
Cat. no.
1021200
Kit size
50
Links
Norgen Biotek Protocol Information Sheet

Features

• Rapid spin-column format allows for the processing of 10 samples in 45 minutes.
• RNA is isolated without the use of harmful chemicals such as phenol or chloroform.
• All leukocyte RNA species can be isolated, from large mRNA species down to microRNA (miRNA).
• Rapid removal of red blood cells from whole blood samples using differential red blood cell lysis.
• Purified RNA 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 analysis requiring the use of intact RNA.

Norgen'’s Leukocyte RNA Purification Kit provides a rapid method for the 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 purified RNA is of the highest quality and can be used in a number of downstream applications.

Purification is based on spin column chromatography using Norgen’s proprietary resin as the separation matrix. The RNA is preferentially purified from the other cellular components such as proteins, without the use of phenol or chloroform. For leukocyte RNA purification, whole blood samples are first collected with anticoagulants. The red blood cells are removed through differential red blood cell lysis, and the leukocytes are recovered by centrifugation (please see flow charts on pages 4 and 5). The recovered leukcoytes are then lysed, and the lysate is loaded onto a supplied spin column. Norgen’s resin binds RNA in a manner that depends on ionic concentrations. Thus only the RNA will bind to the column, while the contaminating proteins will be removed in the flowthrough or retained on the top of the resin. The bound RNA is then washed three times with the provided wash solution in order to remove any remaining impurities, and the purified leukocyte RNA is eluted with the elution buffer. Norgen’s kit allows for the isolation of total leukocyte RNA, including all small RNA species. 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.

Column Binding Capacity	50 µg
Size of RNA purified	All sizes
Minimum blood input	10 µl
Maximum blood input	2 ml or 3x10exp6 leukocytes
Maximum Column Loading volume	600 µl
Time to complete 10 purifications	45 min
Average RNA yield	500 µl Human blood: 1.5 µg

Leukocyte total RNA:

Leukocyte total RNA was isolated from 250 µL of rabbit blood from 4 different samples (lanes 1 to 4). Twenty µL of each elution was then run on a 1.5% formaldehyde-agarose gel. Lane M is Norgen's RNA Ladder.

Consistency and Yield vs. Competitor:

Leukocyte total RNA was isolated from 50 µL of human blood. Twenty uL out of 50 µL of each elution was then run on a 1.5% formaldehyde-agarose gel. Lane M is Norgen's RNA Ladder. Note the quality, consistency and yield of Norgen's kit (Lane 1 to 4) as compared to the competitor (Lane 5 and 6).

Procedure Diagram