rabbit anti rabin8  (Proteintech)

Journal: iScience

Article Title: Protein quality control machinery supports primary ciliogenesis by eliminating GDP-bound Rab8-family GTPases

doi: 10.1016/j.isci.2023.106652

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Article Snippet: anti-RABIN8/RAB3IP rabbit polyclonal , Proteintech , Cat# 12321-1-AP; RRID: AB_2177510.

Techniques: Recombinant, Protease Inhibitor, Modification, Marker, Membrane, Diagnostic Assay, Transfection, Plasmid Preparation, DNA Extraction, Sequencing, Negative Control, Software

Journal: Developmental cell

Article Title: Akt regulates a Rab11-effector switch required for ciliogenesis

doi: 10.1016/j.devcel.2019.05.022

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Rabbit polyclonal anti-RAB3IP/Rabin8 , Proteintech , 12321-1-AP, RRID:AB_2177510.

Techniques: Immunofluorescence, Virus, Recombinant, Sequencing, Software

Journal: The Journal of Biological Chemistry

Article Title: Golgi-resident TRIO regulates membrane trafficking during neurite outgrowth

doi: 10.1074/jbc.RA118.007318

Figure Lengend Snippet: TRIO promotes RABIN8 phosphorylation and regulates RABIN8 activity. A, RABIN8 phosphorylation analysis in cerebella tissues. B, quantification of the relative level of phosphorylated RABIN8 normalized to the pelleted RABIN8 in the blots shown in A. The error bars indicate S.E. (Student's t test). *, p < 0.05; n = 3. C, RABIN8 phosphorylation analysis in CGNs. D, quantification of RABIN8 phosphorylation in C. The error bars indicate S.E. (Student's t test). *, p < 0.05; n = 3. E, P10 mouse cerebellum was homogenized, and the postnuclear supernatants were subjected to 2.5–30% OptiPrep density gradient for subcellular fractionation. Fractions were subjected to Western blotting with RABIN8, RAB8, RAB10, and ERK1/2 antibodies. F, CGNs were cultured for 2 DIV and subjected to immunofluorescence using RABIN8 and RAB8 antibodies. The scale bar represents 5 μm. Scatter plots and the PCC of the fluorescence intensities of red and green channels were also shown. G, quantification of RABIN8 and RAB8 co-localization in neurites of CGNs as shown in F. Pearson's correlation coefficient and Manders' coefficient M1 and M2 were analyzed. The error bars indicate S.E. (Student's t test). *, p < 0.05. Each group comprised three mice (n = 3) and 18 neurons/group. A.U., arbitrary units; n.s., not significant; IP, immunoprecipitation; MW, molecular weight.

Article Snippet: Lysates were cleared by centrifugation at 12000 rpm for 10 min; some cleared supernatants were retained as the total input, and the remaining lysates were incubated with 20 μl of a 50% slurry of Anti-FLAG M2 affinity gel (A2220, Sigma–Aldrich) or 40 μl of a 50% slurry of protein A–Sephorose (catalog no. 17-0974-01, GE Healthcare) plus the rabbit anti-TRIO antibody (H120, sc-28564, Santa Cruz Biotechnology), the rabbit anti-RABIN8 antibody (12321-1-AP, Proteintech), or equal amount of the normal rabbit IgG (sc-2027, Santa Cruz Biotechnology) overnight at 4 °C.

Techniques: Phospho-proteomics, Activity Assay, Fractionation, Western Blot, Cell Culture, Immunofluorescence, Fluorescence, Immunoprecipitation, Molecular Weight

Journal: The Journal of Biological Chemistry

Article Title: Golgi-resident TRIO regulates membrane trafficking during neurite outgrowth

doi: 10.1074/jbc.RA118.007318

Figure Lengend Snippet: TRIO directly interacts with RABIN8. A, co-immunoprecipitation of endogenous TRIO and RABIN8 in P21 mouse cerebellum. B, co-immunoprecipitation of EGFP-TRIO8 and FLAG-RABIN8 in Neuro-2a cells. C, co-immunoprecipitation of FLAG-RABIN8 with EGFP-TRIO (1–1295) and TRIO (1296–1909) in COS-7 cells. D, GST-RABIN8 pulldown assay of P21 mouse cerebellum lysates. E, RABIN8 phosphorylation assay to CGNs treated with ITX3. RAC1 activity was decreased. F, quantification of RABIN8 phosphorylation in E. The error bars indicate S.E. (Student's t test); n = 3. G, direct binding assay of His6-RABIN8 to GST–TRIO variants. Coomassie Blue staining of GST–TRIO variants are shown at the bottom. H, COS-7 cells were transfected with FLAG-RABIN8, together with either pEGFP-C3, pEGFP-TRIO(1–230), pEGFP-TRIO(208–673), pEGFP-TRIO(446–909), pEGFP-TRIO(672–1295), or pEGFP-TRIO(1296–1909), and were lysed and subjected to immunoprecipitation. RABIN8 phosphorylation level was determined. I, quantification of the pRABIN8 level in H. The error bars indicate S.E. (one-way ANOVA with Bonferroni's test). *, p < 0.05; **, p < 0.01; n = 6. n.s., not significant; A.U., arbitrary units; IP, immunoprecipitation. MW, molecular weight; WB, Western blot.

Article Snippet: Lysates were cleared by centrifugation at 12000 rpm for 10 min; some cleared supernatants were retained as the total input, and the remaining lysates were incubated with 20 μl of a 50% slurry of Anti-FLAG M2 affinity gel (A2220, Sigma–Aldrich) or 40 μl of a 50% slurry of protein A–Sephorose (catalog no. 17-0974-01, GE Healthcare) plus the rabbit anti-TRIO antibody (H120, sc-28564, Santa Cruz Biotechnology), the rabbit anti-RABIN8 antibody (12321-1-AP, Proteintech), or equal amount of the normal rabbit IgG (sc-2027, Santa Cruz Biotechnology) overnight at 4 °C.

Techniques: Immunoprecipitation, Phospho-proteomics, Activity Assay, Binding Assay, Staining, Transfection, Molecular Weight, Western Blot