This antibody gave a positive signal in HeLa whole cell lysate and HeLa nuclear lysate in Western blot, HeLa whole cell lysate in Immunofluorescence and Human Kidney tissue lysate in Immunohistochemistry.
Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C or -80°C. Avoid freeze / thaw cycle.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use a concentration of 1 µg/ml.
Use a concentration of 1 µg/ml.
Use a concentration of 1 µg/ml. Detects a band of approximately 75 kDa (predicted molecular weight: 74 kDa).
Abcam recommends using milk as the blocking agent.
The protein encoded by this gene is a member of the RecQ DNA helicase family. DNA helicases are enzymes involved in various types of DNA repair, including mismatch repair, nucleotide excision repair and direct repair. Some members of this family are associated with genetic disorders with predisposition to malignancy and chromosomal instability.
This blot was produced using a 4-12% Bis-tris gel under the MOPS buffer system. The gel was run at 200V for 50 minutes before being transferred onto a Nitrocellulose membrane at 30V for 70 minutes. The membrane was then blocked for an hour using 3% Milk before being incubated with ab22830 overnight at 4°C. Antibody binding was detected using an anti-rabbit antibody conjugated to HRP, and visualised using ECL development solution ab133406.
ICC/IF image of ab22830 stained human HeLa cells. The cells were methanol fixed (5 min) and incubated with the antibody (ab22830, 1µg/ml) for 1h at room temperature. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Image-iTTM FX Signal Enhancer was used as the primary blocking agent, 5% BSA (in TBS-T) was used for all other blocking steps. DAPI was used to stain the cell nuclei (blue). Alexa Fluor® 594 WGA was used to label plasma membranes (red).
Panel A shows localisation of ab22830 to the nuclei, Panel B has the Alexa Fluor® 488 channel removed for comparison.
IHC image of ab22830 staining RECQ1 in Human kidney formalin fixed paraffin embedded tissue section, performed on a Leica BondTM system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab22830, 1µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.
For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.
Western blot - Anti-RECQ1 antibody (ab22830)
All lanes : Anti-RECQ1 antibody (ab22830) at 1 µg/ml
Lane 1 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate at 20 ug Lane 2 : HeLa (Human epithelial carcinoma cell line) Nuclear Lysate at 20 ug
Lane 3 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate at 20 ug with Human RECQ1 peptide (ab25318) at 1 µg/ml Lane 4 : HeLa (Human epithelial carcinoma cell line) Nuclear Lysate at 20 ug with Human RECQ1 peptide (ab25318) at 1 µg/ml
Secondary All lanes : Goat polyclonal to Rabbit IgG (Alexa Fluor® 680) at 1/10000 dilution
ab22830 recognizes a band at 75 kDa which corresponds in size to that of RECQ1. Several smaller bands are also detected which may likely correspond to degradation products. All these bands are almost entirely competed away by the addition of the immunizing peptide, suggesting tha the interaction is specific.