Product nameHuman NEFM (160 kD Neurofilament Medium) knockout HEK293T cell lysate
Knockout cell lysate achieved by CRISPR/Cas9.
Parental Cell LineHEK293T
Mutation descriptionKnockout achieved by using CRISPR/Cas9, 1 bp insertion in exon 2 and Insertion of the selection cassette in exon 2.
Knockout validationSanger Sequencing, Western Blot (WB)
Reconstitution notesTo use as WB control, resuspend the lyophilizate in 50 µL of LDS* Sample Buffer to have a final concentration of 2 mg/ml. For reducing conditions, we recommend a final concentration of 0.1 M DTT.
*Usage of SDS sample buffer is not recommended with these lyophilized lysates.
Lysate preparation: Our lysates are made using RIPA buffer to which we add a protease inhibitor cocktail and phosphatase inhibitor cocktail (ratio: 300:100:10). This means that the protein of interest is denatured. If you require a native form of the protein please use the live cell version - found here. Please refer to our lysis protocol for further details on how our lysates are prepared.
User storage instructions: After reconstitution, store the lysate at -80°C.
Access thousands of knockout cell lysates, generated from commonly used cancer cell lines.
See here for more information on knockout cell lysates.
Abcam has not and does not intend to apply for the REACH Authorisation of customers’ uses of products that contain European Authorisation list (Annex XIV) substances.
It is the responsibility of our customers to check the necessity of application of REACH Authorisation, and any other relevant authorisations, for their intended uses.
This product is subject to limited use licenses from The Broad Institute, ERS Genomics Limited and Sigma-Aldrich Co. LLC, and is developed with patented technology. For full details of the licenses and patents please refer to our limited use license and patent pages.
Tested applicationsSuitable for: WBmore details
Storage instructionsStore at -80°C. Please refer to protocols.
Components 1 kit ab260195 - Human NEFM knockout HEK293T cell lysate (Lyophilized) 1 x 100µg ab255553 - Human wild-type HEK293T cell lysate (Lyophilized) 1 x 100µg
STR AnalysisAmelogenin X D5S818: 8, 9 D13S317: 12, 14 D7S820: 11 D16S539: 9, 13 vWA: 16, 19 TH01: 7, 9.3 TPOX: 11 CSF1PO: 11, 12
RelevanceNeurofilaments are the 10nm or intermediate filament proteins found specifically in neurons, and are composed predominantly of three major proteins called neurofilament light (NF-L), neurofilament medium (NF-M) and neurofilament heavy (NF-H). Neurofilament medium runs on SDS-PAGE gels in the range 145-170 kDa, with some variation in different species. Antibodies to this protein are useful to identify neurons and their processes in tissue sections and in tissue culture. Neurofilament medium can also be useful in studies of neurofilament accumulations seen in many neurological diseases, such as Lou Gehrig's disease or Alzheimer's disease.
- 150kDa medium
KO cell lines
Our Abpromise guarantee covers the use of ab257103 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|WB||Use at an assay dependent concentration. Predicted molecular weight: 102 kDa.|
Lane 1:Wild-type HEK-293T (Human epithelial cell line from embryonic kidney transformed with large T antigen) whole cell lysate (20 ug)
Lane 2:NEFM knockout HEK-293T (Human epithelial cell line from embryonic kidney transformed with large T antigen) whole cell lysate (20 ug)
Lane 3:A549 (Human lung carcinoma cell line) whole cell lysate (20 ug)
ab7794 was shown to specifically react with 160 kD Neurofilament Medium in wild-type HEK-293T cells. Loss of signal was observed when knockout cell line ab266741 (knockout cell lysate ab257103) was used. Wild-type and 160 kD Neurofilament Medium knockout samples were subjected to SDS-PAGE. ab7794 and Anti-GAPDH antibody[EPR16891] - Loading Control (ab181602) were incubated overnight at 4oC at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 680RD) preadsorbed (ab216777) and Goat anti-Mouse IgG H&L (IRDye® 800CW) preadsorbed (ab216772) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.
Allele-1: 1 bp insertion in exon 2
Allele-2: Insertion of the selection cassette in exon 2
To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.
ab257103 has not yet been referenced specifically in any publications.