Immunofluorescence analysis of Phospho-Hsp25 pSer86 was done on 70% confluent log phase HeLa cells treated with UV-302nm for 60 minutes. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 10 minutes, and blocked with 1% BSA for 1 hour at room temperature. The cells were labeled with Phospho-Hsp25 pSer86 Rabbit Polyclonal Antibody (44536G) at 2ug/ml in 0.1% BSA and incubated for 3 hours at room temperature and then labeled with Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A27034) at a dilution of 1:2000 for 45 minutes at room temperature (Panel a: green). Nuclei (Panel b: blue) were stained with SlowFade® Gold Antifade Mountant with DAPI (S36938). F-actin (Panel c: red) was stained with Alexa Fluor® 555 Rhodamine Phalloidin (Product # R415, 1:300). Panel d is a merged image showing cytoplasmic localization. Panel e is untreated cell with no signal. Panel f is a no primary antibody control. The images were captured at 60X magnification.
|Tested species reactivity||Human, Mouse|
|Published species reactivity||Mouse|
|Host / Isotype||Rabbit / IgG|
|Immunogen||The antiserum was produced against a chemically synthesized phosphopeptide derived from a region of mouse HSP25 that contains serine 86.|
|Purification||Antigen affinity chromatography|
|Storage buffer||Dulbecco's PBS, pH 7.3, with 1mg/ml BSA, 50% glycerol|
|Contains||0.05% sodium azide|
|Tested Applications||Dilution *|
|Flow Cytometry (Flow)||3-5 µg/10^6 cells|
|Immunocytochemistry (ICC)||2-3 µg/mL|
|Immunofluorescence (IF)||2-3 µg/mL|
|Western Blot (WB)||1:1000|
* Suggested working dilutions are given as a guide only. It is recommended that the user titrate the product for use in their own experiment using appropriate negative and positive controls.
|Western Blot (WB)||See 1 publications below|
HSP27, also known as Heat shock 27 kDa protein, is a cytoplasmic protein in interphase cells and colocalizes with mitotic spindles in mitotic cells. It translocates to the nucleus during heat shock. HSP27 belongs to the small heat shock protein (HSP20) family and is known to be an important protein in stress resistance and actin organization and associates with alpha- and beta-tubulin, microtubules and CRYAB. It also interacts with other heat shock proteins like HSPB8 and HSPBAP1. HSP27 is expressed in response to environmental stresses such as heat shock, or estrogen stimulation in MCF-7 cells. Ubiquitous expression is seen in most tissues with highest levels in heart and in tissues composed of striated and smooth muscle. Defects in HSPB1 are the cause of Charcot-Marie-Tooth disease type 2F (CMT2F) and distal hereditary motor neuronopathy type 2B (HMN2B).
For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.
|Mouse||Not Cited||p38 MAPK/HSP25 signaling mediates cadmium-induced contraction of mesangial cells and renal glomeruli.||Hirano S,Sun X,DeGuzman CA,Ransom RF,McLeish KR,Smoyer WE,Shelden EA,Welsh MJ,Benndorf R||American journal of physiology. Renal physiology (288:F1133)||2005|