The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use at an assay dependent concentration.
Use a concentration of 1 µg/ml. Detects a band of approximately 85, 140 kDa (predicted molecular weight: 66, 113 kDa).
Use a concentration of 0.5 - 5 µg/ml. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
Use a concentration of 1 µg/ml.
Probable transcription factor.
Expressed in testis.
Defects in SALL4 are the cause of Duane-radial ray syndrome (DRRS) [MIM:607323]; also known as Okihiro syndrome. DRRS is a disorder characterized by the association of forearm malformations with Duane retraction syndrome. Defects in SALL4 are the cause of oculootoradial syndrome (OORS) [MIM:147750]. Oculootoradial syndrome is an autosomal dominant condition characterized by upper limbs anomalies (radial ray defects, carpal bones fusion), extraocular motor disturbances, congenital bilateral non-progressive mixed hearing loss. Other less consistent malformations include heart involvement, mild thrombocytopenia and leukocytosis (before age 50), shoulder girdle hypoplasia, imperforate anus, kidney malrotation or rectovaginal fistula. The IVIC syndrome is an allelic disorder of Duane-radial ray syndrome (DRRS) with a similar phenotype.
Belongs to the sal C2H2-type zinc-finger protein family. Contains 7 C2H2-type zinc fingers.
IHC image of ab29112 staining in human testis 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 ab29112, 5µg/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.
*Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre.
Immunocytochemistry/ Immunofluorescence - Anti-Sall4 antibody (ab29112)This image is courtesy of an anonymous abreview.
ICC/IF image of ab29112 stained human embryonic stem cells. The cells were fixed in Paraformaldehyde, permeabilized using 0.1% Triton X-100, blocked with 10% Goat serum, 0.1% BSA in PBS for 1 hour at RT, before incubation with ab29112 at a 1/100 dilution for 12 hours at 4°C. The secondary used was an Alexa Fluor® 488 conjugated goat anti-rabbit polyclonal, at 1/200 dilution.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Sall4 antibody (ab29112)This image is courtesy of an abreview by Zachary Yu-Ching Lin.
IHC-P image of Sall4 staining with ab29112 on tissue sections from a 3 year old adult marmoset testis. The sections were subjected to heat-mediated antigen retrieval using Dako antigen retrieval solution. The sections were then blocked with 5% milk for 30 minutes at 25°C, before incubation with ab29112 (1/100 dilution) for 20 hours at 4°C. The secondary was an Alexa Fluor® 555 conjugated goat anti-rabbit polyclonal, used at a 1/500 dilution.
Secondary IR Dye 680 Conjugated Goat Anti-Rabbit IgG (H+L) at 1/15000 dilution
Performed under reducing conditions.
Predicted band size : 66, 113 kDa Observed band size : 85,140 kDa (why is the actual band size different from the predicted?) This antibody detects two bands which we believe correspond to the two isoforms of Sall4 outlined in Ma et al, 2006 (PMID: 16763212). This paper describes human isoforms running at 165 kDa and 95 kDa. In mouse embryonic stem cells we see both isoforms running at slightly lower molecular weights. We believe that these bands represent Sall4a (Swissprot: Q8BX22) and Sall4b (Swissprot: Q6S7E9) which are predicted to be 113 kDa and 66 kDa respectively.
ICC/IF image of ab29112 stained mouse embryonic stem cells. The cells were 4% PFA fixed (10 min) and then incubated in 1% BSA/10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab29112, 1µg/ml) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue).