Some assays require direct immobilization of a histidine tagged protein, in which case nickel or coppercoated plates are suitable these plates can also be used. The plate, which comprises of 8 wells x 12 strips, is demountable. Histag elisa for the detection of humoral tumorspecific. Nta, which has four chelating sites for nickel ions, binds nickel more tightly than. Human glutathione peroxidase 1 gpx1 protein his tag,t7. Antihis tag antibody mouse monoclonal hrp elisa,ihc,wb lsbio. Aviscera bioscience offers fgf21 elisa kits enables to detect human, mouse or rat samples. Antihis tag antibody rabbit polyclonal elisa,ihc,wb lsbio. Specific detection of histagged proteins with recombinant anti. Enzymelinked immunosorbent assay elisa introduction in addition to western blots, the anti hisc term antibodies can also be used in immunoassays. Antigen immobilization antigen immobilization varies between two principle techniques.
His protein elisa standard this kit contains one his protein elisa standard item no. Biovisions histag protein elisa kit is a competitivebased elisa that is easier, faster and more sensitive method to detect his tag proteins expressed in both bacterial, insect and mammalian cells. The antibody recognizes the 6xhis tag fused to either the amino or carboxy terminus of targeted proteins in transfected cells. It is a sandwich elisa kit where the recombinant proteins his gst tags are captured with one tag and detected with the other tag. Browse a collection of his tagged protein purification protocols learn about our quick and easy protein purification kit protocols. Sensitivity range of the kit is 4 g ml to 1 ng ml, or 950 nm to 50 pm of 6xhis tag residues. Per liter of culture, centrifuge cells and resuspend in 50 ml of his binding. Histag recombionat gsthis tag fusion protein elisa kit. Detection and assay using the qiaexpress tag100 antibody. Arg81201 6x his tagged protein elisa kit is an enzyme immunoassay kit for the quantification of 6x his tagged proteins or residues in cell or tissue lysates. Pdf his tag elisa for the detection of humoral tumor. Propur affinity spin columns contain a unique flow regulator, flowgo, which maximizes the contact time between the target protein and the resin. Elisa technical guide and protocols thermo fisher scientific.
Standard curves must be run each time an assay is performed. Mouse monoclonal antibody for the detection of his6tagged recombinant proteins. The elisa enzymelinked immunosorbent assay is a widely used application for detecting and quantifying proteins and antigens. The optimized cobaltbased immobilized metal affinity.
Recombinant proteins with his tag on its n or c terminus. Protein purification protocols his tag agarose cube biotech. His tag horseradish peroxidaseconjugated antibody mab050h. Histag antibody detects recombinant proteins containing the 6xhis epitope tag. Direct detection of histagged proteins using nickelnta. This detection kit offers readytouse reagents, and can detect as low as 20 ngml of his tag protein within 90 min. The antibody recognizes the his tag fused either to the amino or carboxy termini of targeted proteins.
This experiment was conducted under reducing conditions and using the 12230 kda separation system. This monoclonal anti6x his tag antibody detects overexpressed proteins containing the 6x his epitope tag and is useful in determining its presence in various assays. This antibody has been tested by elisa and western blotting against both the immunizing peptide and his containing recombinant proteins. Elisa kit where the recombinant proteins hisgst tags are captured with one tag and detected with the other tag. The antibody may crossreact with a protein of unknown origin 6070kda. Sample concordance between capture and indirect assays was highly. Aug 14, 2012 the elisa kti contains two antibodies. Applications invitrogen histag antibodies are validated for use in applications such as elisa, western blot, flow cytometry, immunofluorescence, immunohistochemistry, immunocytochemistry, and immunoprecipitation. Alternatively, precoated avidinstreptavidin plates can be used. The enzyme linked immunosorbent assay elisa is a powerful method for detecting and quantifying a specific protein in a complex mixture. Pdf histag elisa for the detection of humoral tumor. Originally described by engvall and perlmann 1971, the method enables analysis of protein samples immobilized in microplate wells using specific antibodies.
Monoclonal 6 his epitope tag, monoclonal 6xhis tag, monoclonal antipolyhistidine, monoclonal hhhhhh epitope tag, monoclonal hexa his tag, monoclonal his tag, monoclonal his6 tag, monoclonal histidine tagged, monoclonal poly his tag. If it is used to purify other proteins, the protocol may have to be modified. Direct elisa is used to detect antigen that is directly bound to a plate for example, on whole cells or viruses while indirect elisa, which usually takes the format of a sandwich elisa, uses a multistep protocol to capture the target protein using a second antibody to detect. This 6xhis tag detection assay kit is an enzyme immunoassay developed for the sensitive detection of 6xhis tagged fusion proteins with a suitable detector. Denaturing the sample by reducing all available disulfide bonds may enhance exposure of buried histidine residues and increase the probability of his tag. Solubility and purification of the recombinant flagellin protein with histag. Wb see our web site product page for detailed applications information protocols. Avisceras anti fgf 21 antibody works well at western blot, immunohistochemistry. Histag protein elisa kit protein quantitation kits. Because of its small size, strep tag generally does not interfere with the biological activity of the fusion partner.
His tag antibody nbp125939 200, 100, or 50 ng of e. After electrophoresis and transfer of the proteins to a appropriate. A sample protocol is provided for your convenience. All purified histidine tagged proteins were immunological active and 90% pure. Sarscov 2, order animal sera against sarscovo2 or antihis tag antibodies the. These are available online in convenient and compact pdf format at. His tag purification protocol university of san diego home pages. His tagged protein purification requires the his tag and ninta interaction, which is based on the selectivity and high affinity of ninta nickel nitrilotriacetic acid resin for proteins containing an affinity tag of e. Protocol the following purification protocol is optimized for the purification of mbpendoglycosidase. His tag segments of proteins may fold in a manner that reduces interaction between the his tag and the nickelnta conjugate. Save your time and cost one hour assay high sensitivity. Oryza sativa ribosomal protein s8 rps8 protein his tag.
The purified material was characterised by sdspage, elisa and western blot. Gtx62891401 wb image 6x his antibody conjugated hrp detects 6x his protein by western blot analysis. To date, this antibody has reacted with all his tagged proteins so far tested. Precision experiments resulted in cvs his tag and indirect elisa accurately identified 88% and 93% of samples, respectively. It is also known as hexa histidine tag, 6xhis tag, his6 tag, by the us trademarked name his tag us trademark serial number 74242707, and most commonly as his tag. Propur columns offer the speed of a spin column with the yields and purities of a gravity column or a. This site lists the protocol to apply for his tag protein purifications using our purecube agarose resins. Download this native purification protocol as a pdf file. Fgf21 is a biomarker of diabetes, obesity and metabolic syndrome. Insect cells recombinant 95 % as determined by sds page, size exclusion chromatography and western blot. Perform western blot experiment using pentahis antibody. As this is a rapid screening assay, and the immunoreactivity of different his tagged proteins is highly variable,2 the standard is not designed to be used as a quantitative tool, but provide a means of monitoring protein gain or loss during the various. As such, anti6x his 6x his can be used to identify fusion proteins containing the 6x his epitope. A polyhistidine tag is an amino acid motif in proteins that typically consists of at least six histidine his residues, often at the n or cterminus of the protein.
For more information, please refer to antibodies harlow. The detector antibody in the kit is rabbit antigst, and this is in turn detected by an antirabbit igg antibody conjugated to hrp. His tag purification purification protocol theoryandintroduction. Production of recombinant flagellin to develop elisabased. His tag binding plates nickel and copper chelatecoated plates are ideal for analyzing polyhistidinetagged fusion proteins by elisa based methods. The interaction of 6xhis tagged proteins with immobilized nickel ions is ph dependent. When anti his monoclonal antibody and his tagged proteins are added to the well, the coated his tagged. Proteins that contain a succession of several histidine residues at the amino or carboxyl terminus have a strong binding affinity for metal. Purification of polyhistagged recombinant proteins.
Protein purification animation his tag protein purification this animation lecture explains about the histidine tag mediated protein purification. Detection of his tag recombinant protein in the samples. In a traditional direct coating elisa, antigens are directly attached to the plate by passive adsorption, usually using a carbonatebicarbonate buffer at ph 9. Mouse monoclonal antibody for the detection of histidinetagged recombinant proteins. His tag gst tag purification and detection tools nunc.
This polyclonal anti6x his tag antibody detects overexpressed proteins containing the 6x his epitope tag. Detection of his tagged recombinant proteins in the samples. His tag plate in the kit is a 96well microtiter plate coated with a his tagged protein with the molecular weight of 12. May 29, 2008 the his tagged capture elisa could be standardized. Glutathione peroxidase 1 gpx1 aa 1201 protein his tag protein gpx1 origin. Histag mouse monoclonal antibody 4e6 sab signalway. Content expression and purification of proteins using 6xhistidine tag 3 content 1 introduction 4 1. Prepare all wells, reagents, standards and samples as described in sections vii and viii respectively. Ni1affinitychromatography usestheabilityof his tobindnickel. Recombinant proteins with his tag on its n or c terminus fast assay. Peroxidase antibody produced in mouse clone his 1, purified immunoglobulin, lyophilized powder synonym. We use cookies to improve your browsing experience and provide meaningful content. Histag 27e8 mouse mab detects recombinant proteins containing the 6xhis epitope tag.
This detection kit offers readytouse reagents, and can detect as low as 20 ngml of his tag. Histag recombionat gsthis tag fusion protein elisa kit, 96. Enzymelinked immunosorbent assay elisa for plasma igg. Antibody is suitable for elisa, immunoblotting and dot blotting assays. In comparison to western blot analysis, his tag and indirect elisa accurately identified 88% and 93% of samples, respectively. Description technical specs detection of his tag recombinant protein in the samples. His 1, monoclonal biological source mouse applications dot blot. The his tag elisa kit employs the quantitatively competitive enzyme immunoassay technique in which poly his tag present in samples or. Purification of polyhistagged recombinant proteins using. Expression and purification of proteins using 6x histidinetag. It is recommended that all standards and samples should be run at least in duplicate. Cell biolabs histag protein elisa kit is a competitive enzyme immunoassay developed for rapid detection and quantitation of his tagged protein samples c or nterminal.
Specific bands were detected for his tag at approximately 32, 65, and 120 kda as indicated using a 1. Recombinant small acidsoluble spore protein tlp tlp protein his tag. Denaturing the sample by reducing all available disulfide bonds may enhance exposure of buried histidine residues and increase the probability of his tag interaction with nickelnta. Invitrogen histag antibodies reliably detect recombinant his tagged proteins and are validated for use in various applications.
The tag was invented by roche, although the use of histidines and. Caymans his tag detection elisa kit is a competitive assay designed for the rapid, semiquantitative screening of cell lysates and affinity column fractions for. Quickly identify the presence of his tagged proteins in samples. Our histag protein elisa kit allows you to detect and quantify his tagged protein samples simply and reliably by comparing your unknown samples to a known recombinant standard. His tag antibody lsc60077 is an unconjugated rabbit polyclonal antibody to his tag. Anti6x his tag antibody gt359 hrp gtx62891401 genetex. Hrpconjugated goat antimouse igg polyclonal at 15000 dilution developed using the ecl technique.
Our anti his tag antibody portfolio has been tested in elisa, flow cytometry, immunofluorescence and immunoprecipitation clone ad1. Validated in wb, elisa, flow cyt and tested in species independent. The most common his tag is the hexahistidine his66 his tag which has a theoretical molecular weight of 0. Nanocaptureligand human ige, vhh, biotinylated introduction. The quantity of his tag in a protein sample is determined by comparing its absorbance with that of a known his tag protein standard curve provided in the kit.
173 808 889 258 1550 733 1029 82 253 953 745 1258 851 1224 647 127 1469 628 1374 1080 128 193 16 881 1308 270 872 914 530 129 593 1493 213 996 1294