Tested Applications
| Positive WB detected in | Recombinant protein protein |
Recommended dilution
| Application | Dilution |
|---|---|
| Western Blot (WB) | WB : 1:5000-1:50000 |
| It is recommended that this reagent should be titrated in each testing system to obtain optimal results. | |
| Sample-dependent, Check data in validation data gallery. | |
Published Applications
| WB | See 124 publications below |
| IF | See 1 publications below |
| IP | See 3 publications below |
| ELISA | See 7 publications below |
| CoIP | See 4 publications below |
Product Information
HRP-66005 targets 6*His, His-Tag in WB, IF, IP, CoIP, ELISA, Blocking assay, Cell treatment applications and shows reactivity with recombinant protein samples.
| Tested Reactivity | recombinant protein |
| Cited Reactivity | human |
| Host / Isotype | Mouse / IgG1 |
| Class | Monoclonal |
| Type | Antibody |
| Immunogen |
6xHis Peptide Predict reactive species |
| Full Name | 6*His, His-Tag |
| Calculated Molecular Weight | 0.84 kDa |
| Gene Symbol | |
| Gene ID (NCBI) | |
| RRID | AB_2857904 |
| Conjugate | HRP |
| Form | Liquid |
| Purification Method | Protein G purification |
| UNIPROT ID | HISTAG |
| Storage Buffer | PBS with 50% glycerol, 0.05% Proclin300, 0.5% BSA, pH 7.3. |
| Storage Conditions | Store at -20°C. Avoid exposure to light. Stable for one year after shipment. Aliquoting is unnecessary for -20oC storage. |
Background Information
Protein tags are a protein or peptide sequences located either on the C- or N- terminal of the target protein. His-tag is often used for affinity purification and binding assays. Expressed. The His-tag antibody is a useful tool for monitoring of the His-tagged proteins and recognizes His-tags placed at N-terminal, C-terminal, and internal regions of fusion proteins expressed in bacteria, insect, and mammalian cells. A His-tag (polyhistidine tag) consists of at least six histidine residues that are located at the N- or C-terminus of recombinant proteins. It is commonly used for affinity purification and protein binding experiments.
The recombinant protein has only 6 histidine residues as a tag. Is this sufficient to be detected by your antibody? Is the His-tag antibody able to detect N-terminal and C-terminal His tags?
The His-tag antibody was raised using a 6-His tag peptide as the immunogen, which is ~1 kDa. Some scientists use more than 6 histidine residues as a tag for recombinant proteins to increase affinity to metal ions. A 6-histidine tag is sufficient for specific recognition by the His-tag antibody. It is able to recognize both N-terminal and C-terminal His tags of recombinant proteins produced in various expression systems, including bacteria, yeasts, insect, and mammalian cells.
After purification, I can detect my protein by western blotting using the His-tag antibody but can also see additional bands running lower. What might they represent?
They may represent cleavage products of the recombinant protein. Unspecific protein cleavage can occur during protein production (e.g., in bacteria) or protein purification post the cell lysis steps. Changing expression conditions (temperature, time, induction methods, medium) and using protease inhibitors during purification helps to minimize protein degradation. Additional steps, such as ion exchange, size exclusion, or chromatography, allow isolation of the full-length product.
Can a His-tag antibody be used in applications other than western blotting?
Yes, this antibody has been successfully used for immunofluorescence (IF), immunoprecipitation (IP), and indirect ELISA analysis.
Protocols
| Product Specific Protocols | |
|---|---|
| WB protocol for HRP 6*His, His-Tag antibody HRP-66005 | Download protocol |
| Standard Protocols | |
|---|---|
| Click here to view our Standard Protocols |
Publications
| Species | Application | Title |
|---|---|---|
Immunity Detection of SARS-CoV-2-Specific Humoral and Cellular Immunity in COVID-19 Convalescent Individuals. | ||
Immunity Decreased Expression of the Host Long-Noncoding RNA-GM Facilitates Viral Escape by Inhibiting the Kinase activity TBK1 via S-glutathionylation. | ||
Nat Commun Phosphoglycerate dehydrogenase activates PKM2 to phosphorylate histone H3T11 and attenuate cellular senescence | ||
Nat Commun The structure of a Plasmodium vivax Tryptophan Rich Antigen domain suggests a lipid binding function for a pan-Plasmodium multi-gene family |
Reviews
The reviews below have been submitted by verified Proteintech customers who received an incentive for providing their feedback.
FH Zaneta (Verified Customer) (03-06-2023) | I used the antibody to detect small amount of 6xhis and 8xhis - tagged proteins from HEK cells and SF9 cells, on WesternBlot. It works perfect, fast and efficient!
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FH Tom (Verified Customer) (02-03-2023) | Works great to detect recombinant protein from bacteria
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FH Javier (Verified Customer) (12-16-2022) | Even though there was an issue with the shipment and when I received the antibody it was at room temperature, it has worked perfectly fine. Because of the delay I have used it at a 1:5000 dilution an incubated it for 1h 30 min at room temperature. I would totally recommend it.
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FH Tom (Verified Customer) (07-12-2021) | Blocked membrane overnight in 5% BSA in TBS. Tried at both 1:1000 and 1:10,000 dilutions in 5% BSA in TBST for 2.5h at room temperature and got great, specific bands for both dilutions.
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FH Apoorv (Verified Customer) (12-15-2019) | Used to detect protein in Western blots
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FH Soo (Verified Customer) (10-24-2019) | we use this antibody to detect recombinant proteins which has his tag. Used it for western and ELISA and Flow , works well for us. It is cheaper than other brand's his tag antibody, another reason we like this product.
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FH Angel (Verified Customer) (09-09-2019) | It is the best antibody I've ever used for quick check of protein expression and size for heterologous protein production and purification.
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FH Ryan (Verified Customer) (02-04-2019) | This His-tag-HRP conjugated antibody is the single best His-tag antibody I have ever used. I have used dilutions from 1:1000 to 1:25,000 with fantastic results. The background with this antibody is minimal and the sensitivity is superb. This image shows four different proteins with both 6xHis and 8xHis-tags. I have used the antibody in milk and 5% BSA. Incubation times have been as short as 1 hour and as long as 24 hours; the results are always the same. Being conjugated, it can cut the time of a typical Western blot down by a few hours. My standard procedure with this antibody is 1:8,000 in 5% BSA for 1 hour with three 10 minute washes and a 5 second exposure using Cell Signaling SignalFire ECL.
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FH Jennifer (Verified Customer) (12-08-2018) | Excellent product. We achieved to see the production of our protein of interest in an accurate way. Some previous secondary antibodies gave us a lot of background; this one works better.
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