Description
Hybrid-RTM Blood RNA is a complete kit with ready-to-use reagent for the isolation of total RNA from up to 0.25 ml whole blood sample. This kit utilizes the lysis method of RiboExTM LS which has a powerful ability of cell-lysis and the purification method based on glassfiber membrane technology. Fast and convenient procedure of Hybrid-RTM Blood RNA takes only 30 minutes for complete preparation of pure RNA.
Whole blood sample is homogenized and lysed in RiboExTM LS, a monophasic solution containing phenol and guanidium salt, which rapidly lyse cells and inactivates nucleases. In conventional methods, the erythrocytes of mammalian blood which does not contain nuclei (and therefore, RNA either) should be removed by pre-treatment such as osmotic lysis for the separation of leukocytes from whole blood. This additional treatments increase the experiment time and the possibility of RNA breakage, followed by decline of RNA-quality.
Hybrid-RTM Blood RNA does not need the additional treatment of blood sample, and whole blood is lysed in RiboExTM LS in just one step. Then addition of chloroform brings about a separation of the lysate into aqueous and organic phases. After phase-separating, DNA and protein remains in the interphase and the organic phase respectively but released RNA exists in the aqueous phase. The aqueous phase is picked and applied to a EzPureTM to eliminate small amount of contaminated DNA and other blood contaminants.
The passed-through is mixed with buffer RB1, RNA binding buffer, and then the mixture is applied to a mini spin column. After a series of washing with buffer RBW and RNW, pure RNA can be eluted by RNase-free water. Hybrid-RTM Blood RNA is suitable for RNA preparation from 0.1 ml to 0.25 ml mammalian whole blood. The typical yield is 3 ug per 0.25 ml whole blood. The purified RNA can be applicable for the isolation of Poly A+ RNA, Northern blotting, dot blotting, in vitro translation, cloning, RT-PCR, RPA and other analytical procedures.
