Expression of natriuretic peptide receptor A in the developing retina of the mouse / 解剖学报

Objective Our purpose is to investigate the expression of natriuretic peptide receptor A (NPR-A) in the retina and to understand the NPR-A’ s functions during the mouse development .Methods Mice eyes were harvested from E16 ( embryonic day 16 ) to P90 ( postnatal day 90 ) . Total of 127 eyes were used in the study . Immunohistochemistries of NPR-A were carried out .Results During development , NPR-A was widely expressed in the retinal neurons .In the outer nuclear layer , NPR-A began to appear in the inner and outer projections of cone and rod cells at P7, but decreased at P14.From P30 afterward, it continued to express weakly .In the inner nuclear layer , NPR-A expressed in the dendrites of bipolar cells weakly from P 7 to adulthood , whereas no expression in horizontal cells .In the ganglion cell layer, NPR-A started highly to express in the ganglion cell bodies at E 16, and in the meantime, in the nerve fiber layer , ganglion cell axons , NPR-A was expressed highly from embryonic to adult .In the inner and outer plexiform layers, NPR-A was highly expressed at P14, but decreased gradually after P30.In addition, NPR-A also widely expressed in the inner protrusions of Müller cells.Conclusion NPR-A participates in the development of the retina , and may be the key molecule in the developing retinal neurons .Moreover, it plays an important regulatory role in the functional activity of Müller cells .

Prokaryotic expression,purification and polyclonal antibody preparation of C terminal Helicase domain of mouse Rig-I / 中国免疫学杂志

Objective:To develop a highly efficacious and sensitive immunological reagent for further investigation on the retinoic acid-induced gene I (Rig-I) of mouse .Methods:The Helicase domain coding region (726-2 240 bp) of mRig-I-H was cloned into plasmid pET15b (+) to construct the recombinant plasmid pET15b(+)-mRig-I-H.Then the plasmid was transformed into E.coli BL21 for protein expression.Rabbits were immunized with electrophoresis-purified recombinant protein to obtain the polyclonal antibody against mRig-I-H.The titer of polyclonal antibody was detected by ELISA and the specificity was identified by Western blot and Immunofluorescence.Results:The recombinant protein was expressed successfully in E.coli.Western blot analysis showed that target protein was expressed with a molecular weight of 40 kD.Titer of the polyclonal antibody was about 1∶1?105 by ELISA assay.With this antibody,we could detect the expression of Rig-I in RAW 264.7 cell line by Western blot and Immunofluorescence.Conclusion:The high level expression of Rig-I Helicase domain is induced in E.coli expressing system.Anti-mRig-I-H polyclonal antibody with high titer and fine specificity could be a novel tool in future investigation of Rig-I.