Antibody was affinity purified using an epitope specific to RbBP5 immobilized on solid support. The epitope recognized by A300-109A maps to a region between residue 500 and the C-terminus (residue 538) of human retinoblastoma binding protein 5 using the numbering given in entry NP_005048.2 (GeneID 5929). Immunoglobulin concentration was determined by extinction coefficient: absorbance at 280 nm of 1.4 equals 1.0 mg of IgG.
RBBP-5, RBQ3, retinoblastoma-binding protein 5, retinoblastoma-binding protein RBQ-3, SWD1, SWD1, Set1c WD40 repeat protein, homolog
between 500 and C-term
2 - 8°C
Tris-citrate/phosphate buffer, pH 7 to 8 containing 0.09% Sodium Azide
Detection of human RbBP5 by western blot. Sample: Whole cell lysate (50 µg for WB and Input; one 10 cm plate for IP) from HEK293T cells. Antibody: Affinity purified rabbit anti-RbBP5 antibody A300-109A used at the indicated concentrations for WB or 10 µg for IP (15% of IP loaded). immunoprecipitatesd RbBP5 was detected by WB using 0.2 mg/ml. Detection: Chemiluminescence with 1 second (WB) or 10 second (IP/WB) exposure.
ChIP-chip scatter plot of anti-RbBP5 (A300-109A) enriched DNA binding sites versus input reference DNA. A. 10 µg of A300-109A was used to immunoprecipitate chromatin from K-562 cells according to Ren et al (Genes Dev. 2002 16: 245-256). immunoprecipitatesd DNA and reference DNA were amplified via ligation-mediated PCR and the products labeled with fluorescent dNTPs. The labeled ChIP and reference DNA were pooled, hybridized to a DNA microarray, and analyzed. Data points below the +3 SD curve (red line) represent significantly enriched binding sites. B. As a control, a similar experiment was performed using normal rabbit IgG. Compared to the anti-RbBP5 ChIP, normal rabbit IgG showed little enrichment.
Localization of RbBP5 Binding Sites by ChIP-sequencing. Chromatin from K562 cells was immunoprecipitated with anti-RbBP5 antibody A300-109A and analyzed by DNA sequencing. The figure illustrates the peak distribution of RbBP5 binding within a 500 Kb region of chromosome 1 as detected using anti-RbBP5 antibody A300-109A. ChIP-seq validation performed by Diogenode, Denville, NJ.