Western Blot: PGP9.5 / UCHL-1 Antibody [NB300-675] - Western blot analysis of extracts from C6 cells using PGP9.5 / UCHL-1 antibody (NB300-675, 1:200). Image from verified customer review.
Immunocytochemistry/ Immunofluorescence: PGP9.5 / UCHL-1 Antibody [NB300-675] - IF Confocal analysis of C6 cells using PGP9.5 / UCHL-1 antibody (NB300-675, 1:5). An Alexa Fluor 488-conjugated Goat to rabbit IgG was used ...read more
This UCHL1 antibody is useful for Immunocytochemistry/Immunofluorescence, Western blot and Immunohistochemistry-Paraffin. By Western blot a band at ~25 kDa is seen.
The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.
25 kDa. Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.
UCH-L1 (ubiquitin carboxyl-terminal hydrolase isozyme L1) was the first discovered de-ubiquitinating enzyme which implicates in processing of ubiquitin precursors and also of ubiquitinated proteins. UCH-L1 is a thiol protease that recognizes and hydrolyzes a peptide bond at C-terminal glycine of ubiquitin. It also binds to free monoubiquitin and prevents its degradation in lysosomes. Localized in cytoplasm and ER membrane as lipid-anchor, UCH-L1 expression is restricted to brain, peripheral nerves, endocrine tissues and gonads of both sexes etc. UCH-L1 deletion in mice leads to fatal neurodegenerative disorder known as gracile axonal dystrophy and it is down-regulated in brains from Parkinson as well as Alzheimer disease patients. Expression outside of neuro-endocrine tissues is found in various cancers including B-cell lymphoma, multiple myeloma, and lung cancer. In transgenic mouse model, UCH-L1 has been demonstrated as an oncogene that causes malignancies by boosting AKT signalling. Furthermore, UCH-L1 has been shown to interfere with ubiquitination of RAPTOR which is catalyzed by DDB1-Cul4 E3 ligase complex, leading to loss of mTORC1 integrity accompanied by a concurrent increase in mTORC2, likely due to increased availability of free mTOR.
This product is for research use only and is not approved for use in humans or in clinical diagnosis. Primary Antibodies are guaranteed for 1 year from date of receipt.
Tested in gray short-tailed opossum (Monodelphis domestica) in testis from 8-month animal. Tissue was fixed in 4% Paraformaldehyde overnight, incubated in 30% sucrose/PBS overnight and embedded in OCT. Sections were cut on a cryostat at 10um.
For immunofluorescence staining: Slides were dried 15 minutes, washed 3x3 min TBST (0.1% Triton X-100), permeabilized for 10 minutes in 2% Triton X-100 in TBS, rinsed 3x3 min TBST, blocked in 10% heat inactivated goat serum in TBST for 30 min, and primary added at 1:100 in 10% blocking solution. Slides were coverslipped with parafilm overnight at 4C. The next day slides were washed with TBST 4x5 min, secondaries (AlexaFluor 568 goat anti rabbit and AlexaFluor 488 goat anti mouse 1:500) for 1 hour, 3x3 washes TBST, and mounted with VectaShield HardSet (+ DAPI).
Intense staining at periphery of seminiferous tubules of Uchl-1 (red in image) with these conditions. 1:200 also worked though staining was less intense (not shown). The same conditions with 1% Triton X-100 for permeabiliziation resulted in weak staining at 1:100 and 1:200. However, ID-4 did not work well under any conditions. Shown in green is some faint staining in interstitial cells but this was not consistent between samples and did not appear to be specific. ID-4 was tested in 4 and 8 month opossum testes, and at dilutions ranging from 1:25 to 1:500. Single-antibody staining was also performed for each antibody and successful in Uchl-1, not ID-4.
IF Confocal analysis of C6 cells using PGP9.5 / UCHL-1 antibody (NB300-675, 1:5). An Alexa Fluor 488-conjugated Goat to rabbit IgG was used as secondary antibody (green). Actin filaments were labeled with Alexa Fluor 568 phalloidin (red). DAPI was used to stain the cell nuclei (blue).
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