DYKDDDDK Tag (FLAG) Recombinant Mouse Monoclonal Antibody [A2-A4-R] - BSA and Azide free
Rmb: 5600
Catalog# HA610054
DYKDDDDK Tag (FLAG) Recombinant Mouse Monoclonal Antibody [A2-A4-R] - BSA and Azide free
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WB
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IP
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IF-Cell
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IHC-P
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Species independent
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HA601167
含抗保成分
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unconjugated
概述
产品名称
DYKDDDDK Tag (FLAG) Recombinant Mouse Monoclonal Antibody [A2-A4-R] - BSA and Azide free
抗体类型
Recombinant Mouse Monoclonal Antibody
免疫原
Synthetic peptide immune sequence is N-DYKDDDDK-C.
种属反应性
Species independent
验证应用
WB, IP, IF-Cell, IHC-P
偶联
unconjugated
克隆号
A2-A4-R
产品特性
形态
Liquid
浓度
存放说明
Store at 2-8℃. Avoid freeze.
存储缓冲液
1*PBS (pH7.4).
亚型
IgG1
纯化方式
Protein A affinity purified.
应用稀释度
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WB
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1:5,000-1:10,000
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IP
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1-2μg/sample
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IF-Cell
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1:1,000-1:5,000
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IHC-P
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1:1,000
靶点
功能
FLAG-tag, or FLAG octapeptide, or FLAG epitope, is a polypeptide protein tag that can be added to a protein using recombinant DNA technology, having the sequence motif DYKDDDDK (where D=aspartic acid, Y=tyrosine, and K=lysine). It is one of the most specific tags and it is an artificial antigen to which specific, high affinity monoclonal antibodies have been developed and hence can be used for protein purification by affinity chromatography and also can be used for locating proteins within living cells. It has been used to separate recombinant, overexpressed protein from wild-type protein expressed by the host organism. It can also be used in the isolation of protein complexes with multiple subunits, because its mild purification procedure tends not to disrupt such complexes. It has been used to obtain proteins of sufficient purity and quality to carry out 3D structure determination by x-ray crystallography. A FLAG-tag can be used in many different assays that require recognition by an antibody. If there is no antibody against a given protein, adding a FLAG-tag to a protein allows the protein to be studied with an antibody against the FLAG sequence. Examples are cellular localization studies by immunofluorescence, immunoprecipitation or detection by SDS PAGE protein electrophoresis and Western blotting. The peptide sequence of the FLAG-tag from the N-terminus to the C-terminus is: DYKDDDDK (1012 Da). Additionally, it may be used in tandem, commonly the 3xFLAG peptide: DYKDHD-G-DYKDHD-I-DYKDDDDK (with the final tag encoding an enterokinase cleavage site). It can be fused to the C-terminus or the N-terminus of a protein, or inserted within a protein. The tyrosine residue in the FLAG-tag can be sulfated, which can affect antibody recognition of the FLAG epitope. The FLAG-tag can be used in conjunction with other affinity tags, for example a polyhistidine tag (His-tag), HA-tag or myc-tag.
背景文献
暂无
别名
DDDDK epitope tag antibody
DDDK antibody
ddk antibody
DYKDDDDK antibody
DYKDDDDK epitope tag antibody
DYKDDDDK tag antibody
ECS epitope tag antibody
ECS tag antibody
Enterokinase Cleavage Site epitope tag antibody
Enterokinase Cleavage Site tag antibody
展开图片
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Western blot analysis of DYKDDDDK Tag (FLAG) on different lysates with Mouse anti-DYKDDDDK Tag (FLAG) antibody (HA610054) at 1/5,000 dilution.
Lane 1: HeLa cell lysate
Lane 2: HeLa transfected with FLAG-tagged Histon H3.1 (N-terminal) cell lysate
Lysates/proteins at 5 µg/Lane.
Predicted band size: 15 kDa
Observed band size: 18 kDa
Exposure time: 1 minute 30 seconds;
4-20% SDS-PAGE gel.
Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (HA610054) at 1/5,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Mouse IgG - HRP Secondary Antibody (HA1006) at 1/50,000 dilution was used for 1 hour at room temperature. -
Western blot analysis of DYKDDDDK Tag (FLAG) on different lysates with Mouse anti-DYKDDDDK Tag (FLAG) antibody (HA610054) at 1/5,000 dilution.
Lane 1: L-929 cell lysate
Lane 2: L-929 transfected with FLAG-tagged CD5 (C-terminal) cell lysate
Lysates/proteins at 5 µg/Lane.
Predicted band size: 65 kDa
Observed band size: 65 kDa
Exposure time: 1 minute 59 seconds;
4-20% SDS-PAGE gel.
Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (HA610054) at 1/5,000 dilution was used in 5% NFDM/TBST at room temperature for 2 hours. Goat Anti-Mouse IgG - HRP Secondary Antibody (HA1006) at 1/50,000 dilution was used for 1 hour at room temperature. -
DYKDDDDK Tag (FLAG) was immunoprecipitated from 0.2 mg 293T transfected with FLAG-tagged PD-L1 (C-terminal) cell lysate with HA610054 at 2 µg/10 µl beads. Western blot was performed from the immunoprecipitate using HA610054 at 1/20,000 dilution. Anti-Mouse IgG for IP, AlpSdAbs® VHH(HRP) (001-100-005) at 1/5,000 dilution was used for 1 hour at room temperature.
Lane 1: 293T transfected with FLAG-tagged PD-L1 (C-terminal) cell lysate (input)
Lane 2: HA610054 IP in 293T transfected with FLAG-tagged PD-L1 (C-terminal) cell lysate
Lane 3: Mouse IgG instead of HA610054 in 293T transfected with FLAG-tagged PD-L1 (C-terminal) cell lysate
Blocking/Dilution buffer: primary antibody dilution (K1803)
Exposure time: 2 seconds; ECL: K1801 -
DYKDDDDK Tag (FLAG) was immunoprecipitated from 0.2 mg 293T transfected with FLAG-tagged MYRF (N-terminal) cell lysate with HA610054 at 2 µg/10 µl beads. Western blot was performed from the immunoprecipitate using HA610054 at 1/20,000 dilution. Anti-Mouse IgG for IP, AlpSdAbs® VHH(HRP) (001-100-005) at 1/5,000 dilution was used for 1 hour at room temperature.
Lane 1: 293T transfected with FLAG-tagged MYRF (N-terminal) cell lysate (input)
Lane 2: HA610054 IP in 293T transfected with FLAG-tagged MYRF (N-terminal) cell lysate
Lane 3: Mouse IgG instead of HA610054 in 293T transfected with FLAG-tagged MYRF (N-terminal) cell lysate
Blocking/Dilution buffer: primary antibody dilution (K1803)
Exposure time: 2 seconds; ECL: K1801 -
Immunohistochemical analysis of paraffin-embedded HeLa transfected with FLAG-tagged Claudin 18.2 (C-terminal) cells with Mouse anti-DYKDDDDK Tag (FLAG) antibody (HA610054) at 1/1,000 dilution.
The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (HA610054) at 1/1,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX. -
Immunohistochemical analysis of paraffin-embedded HeLa transfected with 2*FLAG-tagged Histone H3.1 (N-terminal) cells with Mouse anti-DYKDDDDK Tag (FLAG) antibody (HA610054) at 1/1,000 dilution.
The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (HA610054) at 1/1,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX. -
Immunocytochemistry analysis of HeLa cells labeling DYKDDDDK Tag (FLAG) with Mouse anti-DYKDDDDK Tag (FLAG) antibody (HA610054) at 1/1,000 dilution.
HeLa cells, transfected with empty control (top, negative) / Flag-tagged Claudin 18.2 (C-terminal) (middle, positive) / 2*Flag-tagged Histon H3.1 (N-terminal) (bottom, positive) expression vector, respectively, were fixed in 4% paraformaldehyde for 15 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 15 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Mouse anti-DYKDDDDK Tag (FLAG) antibody (HA610054) at 1/1,000 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.
DYKDDDDK Tag (FLAG) (HA724169, green) was stained at 1/1,000 dilution overnight at +4℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution.
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