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8339
HER/ErbB Family Antibody Sampler Kit

HER/ErbB Family Antibody Sampler Kit #8339

Western Blotting Image 1

Western blot analysis of extracts from A-431, BxPC3 and HeLa cells using EGF Receptor (D38B1) XP® Rabbit mAb.

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Western Blotting Image 2

Western blot analysis of extracts from SK-BR-3 and MCF7 cells using HER2/ErbB2 (D8F12) XP® Rabbit mAb.

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IHC-P (paraffin) Image 3

Immunohistochemical analysis of paraffin-embedded non-small cell lung carcinoma using HER3/ErbB3 (D22C5) XP® Rabbit mAb.

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Western Blotting Image 4

Western blot analysis of extracts from MCF7 (HER3+) and MDA-MB-231 (HER3-) cells using HER3/ErbB3 (D22C5) XP® Rabbit mAb (upper) or β-Actin (D6A8) Rabbit mAb #8457 (lower).

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Western Blotting Image 5

Western blot analysis of extracts from various cell lines, using HER4/ErbB4 (111B2) Rabbit mAb .

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Western Blotting Image 6

Western blot analysis of extracts of BxPC-3 cells, untreated or EGF-stimulated, using Phospho-EGF Receptor (Tyr1068) (D7A5) XP® Rabbit mAb (upper) and EGF Receptor Antibody #2232 (lower).

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Western Blotting Image 7

Western blot analysis of extracts from cells expressing different activated tyrosine kinase proteins, using Phospho-HER2/ErbB2 (Tyr1221/1222) (6B12) Rabbit mAb (upper) or Phospho-Tyrosine mAb (P-Tyr-100) #9411 (lower). Phospho-HER2/ErbB2 (Tyr1221/1222) (6B12) Rabbit mAb specifically detects phosphorylated HER2/ErbB2 but not other phosphorylated tyrosine kinases.

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Western Blotting Image 8

Western blot analysis of extracts from T-47D cells (serum starved overnight), untreated (-) or treated with Human Neuregulin-1 (hNRG-1) #5218 (100 ng/ml, 15 min; +), using Phospho-HER3/ErbB3 (Tyr1289) (D1B5) Rabbit mAb (upper) or HER3/ErbB3 (1B2E) Rabbit mAb #4754 (lower).

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Western Blotting Image 9

Western blot analysis of extracts from various cell lines expressing different activated protein tyrosine kinases, using Phospho-HER4/ErbB4 (Tyr1284) (21A9) Rabbit mAb or Phospho-Tyrosine Mouse mAb (P-Tyr-100) #9411 (lower).

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Western Blotting Image 10

After the primary antibody is bound to the target protein, a complex with HRP-linked secondary antibody is formed. The LumiGLO® is added and emits light during enzyme catalyzed decomposition.

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IHC-P (paraffin) Image 11

Immunohistochemical analysis of paraffin-embedded MDA-MB-468 (amplified EGFR, left), HT-29 (low EGFR, middle) and CAMA-1 (EGFR negative, right) cells using EGF Receptor (D38B1) XP® Rabbit mAb.

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IHC-P (paraffin) Image 12

Immunohistochemical analysis of paraffin-embedded human breast carcinoma using HER2/ErbB2 (D8F12) XP® Rabbit mAb.

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IHC-P (paraffin) Image 13

Immunohistochemical analysis of paraffin-embedded breast carcinoma using HER3/ErbB3 (D22C5) XP® Rabbit mAb.

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IHC-P (paraffin) Image 14

Immunohistochemical analysis using Phospho-EGF Receptor (Tyr1068) (D7A5) XP® Rabbit mAb on SignalSlide™ Phospho-EGF Receptor IHC Controls #8102 (paraffin-embedded KYSE450 cell pellets, untreated (left) or EGF-treated (right)).

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IHC-P (paraffin) Image 15

Immunohistochemical analysis of paraffin-embedded human breast carcinoma, showing membrane localization, using Phospho-HER2/ErbB2 (Tyr1221/1222) (6B12) Rabbit mAb.

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IHC-P (paraffin) Image 16

Immunohistochemical analysis of paraffin-embedded KYSE-450 cell pellets, untreated (left) or treated with Human Epidermal Growth Factor (hEGF) #8916 (right), using Phospho-EGF Receptor (Tyr1068) (D7A5) XP® Rabbit mAb #3777 (upper) or Phospho-HER3/ErbB3 (Tyr1289) (D1B5) Rabbit mAb (lower). Note lack of reactivity with Phospho-EGFR.

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IHC-P (paraffin) Image 17

Immunohistochemical analysis of paraffin-embedded human hepatocellular carcinoma using EGF Receptor (D38B1) XP® Rabbit mAb.

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IHC-P (paraffin) Image 18

Immunohistochemical analysis of paraffin-embedded SK-BR-3 (Her2 high, left) and MCF7 cell pellets (Her2 low, right) using HER2/ErbB2 (D8F12) XP® Rabbit mAb.

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IHC-P (paraffin) Image 19

Immunohistochemical analysis of paraffin-embedded 293T cell pellets transfected with human erbB family members HER3, EGFR, HER2 and HER4 (from left to right as indicated) using HER3/ErbB3 (D22C5) XP® Rabbit mAb (top panels). Transfections were confirmed using EGF Receptor (D38B1) XP® Rabbit mAb #4267 (lower left), HER2/ErbB2 (D8F12) XP® Rabbit mAb #4290 (lower middle) and HER4/ErbB4 (111B2) Rabbit mAb #4795 (lower right).

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IHC-P (paraffin) Image 20

Immunohistochemical analysis of paraffin-embedded HCC827 xenograft, control (left) or λ phosphatase-treated (right), using Phospho-EGF Receptor (Tyr1068) (D7A5) XP® Rabbit mAb.

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IHC-P (paraffin) Image 21

Immunohistochemical analysis of paraffin-embedded SkBr3 cell pellets untreated (left) EGF-treated (right) either untreated (top) or lambda-phosphatase-treated (bottom), using Phospho-HER2/ErbB2 (Tyr1221/1222) (6B12) Rabbit mAb.

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IHC-P (paraffin) Image 22

Immunohistochemical analysis of parafin-embedded human lung carcinoma using Phospho-HER3/ErbB3 (Tyr1289) (D1B5) Rabbit mAb after retreival with citrate (left) or EDTA (right).

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IHC-P (paraffin) Image 23

Immunohistochemical analysis of paraffin-embedded human placenta using EGF Receptor (D38B1) XP® Rabbit mAb.

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Flow Cytometry Image 24

Flow cytometric analysis of A549 cells, untreated (blue) or EGF-treated (green), using Phospho-EGF Receptor (Tyr1068) (D7A5) XP® Rabbit mAb compared to concentration matched XP® Rabbit (DA1E) mAb IgG Isotype Control #3900 (red).

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IHC-P (paraffin) Image 25

Immunohistochemical analysis of paraffin-embedded human renal adenocarcinoma, using Phospho-HER2/ErbB2 (Tyr1221/1222) (6B12) Rabbit mAb in the presence of control peptide (left) or Phospho-HER2/ErbB2 (Tyr1221/1222) Blocking Peptide #1254 (right).

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IHC-P (paraffin) Image 26

Immunohistochemical analysis of paraffin-embedded human lung carcinoma using Phospho-HER3/ErbB3 (Tyr1289) (D1B5) Rabbit mAb.

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IHC-P (paraffin) Image 27

Immunohistochemical analysis of paraffin-embedded human lung carcinoma using EGF Receptor (D38B1) XP® Rabbit mAb.

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IHC-P (paraffin) Image 28

Immunohistochemical analysis of paraffin-embedded ovarian serous adenocarcinoma using HER3/ErbB3 (D22C5) XP® Rabbit mAb.

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IF-IC Image 29

Confocal immunofluorescent analysis of HeLa cells, untreated (left) or EGF-treated (right), using Phospho-EGF Receptor (Tyr1068) (D7A5) XP® Rabbit mAb (green). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).

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IHC-P (paraffin) Image 30

Immunohistochemical analysis of paraffin-embedded human NCI-H358 xenograft, using Phospho-HER2/ErbB2 (Tyr1221/1222) (6B12) Rabbit mAb.

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IHC-P (paraffin) Image 31

Immunohistochemical analysis of paraffin-embedded MCF7 cell pellets, untreated (left) or treated with hNRG-1 #5218 (right), using Phospho-HER3/ErbB3 (Tyr1289) (D1B5) Rabbit mAb.

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Flow Cytometry Image 32

Flow cytometric analysis of Jurkat cells (red) and Kyse70 cells (blue), using EGF Receptor (D38B1) XP® Rabbit mAb.

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IHC-P (paraffin) Image 33

Immunohistochemical analysis of paraffin-embedded MCF7 (left) or MDA-MB-231 (right) cell pellets using HER3/ErbB3 (D22C5) XP® Rabbit mAb.

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IHC-F (frozen) Image 34

Immunohistochemical analysis of frozen HCC827 xenograft, using Phospho-Her2/ErbB2 (Tyr1221/1222)(6B12) Rabbit mAb.

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IHC-P (paraffin) Image 35

Immunohistochemical analysis of paraffin-embedded HCC827 xenograft, untreated (left) or λ phosphatase-treated (right), using Phospho-HER3/ErbB3 (Tyr1289) (D1B5) Rabbit mAb.

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IF-IC Image 36

Confocal immunofluorescent analysis of A549 cells, untreated (left) or treated with human epidermal growth factor (right), using EGF Receptor (D38B1) XP® Rabbit mAb (green). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).

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Flow Cytometry Image 37

Flow cytometric analysis of MCF7 cells using HER3/ErbB3 (D22C5) XP® Rabbit mAb (blue) compared to Rabbit (DA1E) mAb IgG XP® Isotype Control #3900. Anti-rabbit IgG (H+L), F(ab')2 fragment (Alexa Fluor® 488 conjugate) #4412 was used as a secondary antibody.

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IF-IC Image 38

Confocal immunofluorescent analysis of MCF7 cells (HER3+, left) and MDA-MB-231 cells (HER-, right) using HER3/ErbB3 (D22C5) XP® Rabbit mAb (green). Blue pseudocolor= DRAQ5® #4084 (fluorescent DNA dye).

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Product Includes Quantity Applications Reactivity MW(kDa) Isotype
EGF Receptor (D38B1) XP® Rabbit mAb 4267 20 µl
  • WB
  • IP
  • IHC
  • IF
  • F
H M Mk 175 Rabbit IgG
HER2/ErbB2 (D8F12) XP® Rabbit mAb 4290 20 µl
  • WB
  • IHC
H M 185 Rabbit IgG
HER3/ErbB3 (D22C5) XP® Rabbit mAb 12708 20 µl
  • WB
  • IP
  • IHC
  • IF
  • F
H M 185 Rabbit IgG
HER4/ErbB4 (111B2) Rabbit mAb 4795 20 µl
  • WB
  • IP
H M 180 Rabbit IgG
Phospho-EGF Receptor (Tyr1068) (D7A5) XP® Rabbit mAb 3777 20 µl
  • WB
  • IHC
  • IF
  • F
H M R Mk 175 Rabbit IgG
Phospho-HER2/ErbB2 (Tyr1221/1222) (6B12) Rabbit mAb 2243 20 µl
  • WB
  • IHC
H M 185 Rabbit IgG
Phospho-HER3/ErbB3 (Tyr1289) (D1B5) Rabbit mAb 2842 20 µl
  • WB
  • IP
  • IHC
H M 185 Rabbit IgG
Phospho-HER4/ErbB4 (Tyr1284) (21A9) Rabbit mAb 4757 20 µl
  • WB
H 180 Rabbit 
Anti-rabbit IgG, HRP-linked Antibody 7074 100 µl
  • WB
Goat 

The HER/ErbB Family Antibody Sampler Kit provides an economical means to evaluate the HER/ErbB Family, including the phosphorylation of EGFR, HER2/ErbB2, HER3/ErbB3, and HER4/ErbB4. The control antibodies to each family member are also included. The kit contains enough antibody to perform two western blot experiments with each primary antibody.

Each antibody in the HER/ErbB Family Antibody Sampler Kit recognizes only its specific target. The antibodies do not cross react with other HER/ErbB family members.

Monoclonal antibodies are produced by immunizing animals with a fusion protein containing the cytoplasmic domain of human EGF receptor, a synthetic peptide corresponding to residues near the amino terminus of human ErbB2 protein, the carboxy terminus of human ErbB3 protein, or the carboxy-terminus of human ErbB4 protein. Activation state monoclonal antibodies are produced by immunizing animals with a synthetic phosphopeptide corresponding to residues surrounding Tyr1068 of human EGF receptor protein, tyrosines 1221/1222 of human ErbB2 protein, Tyr1289 of human HER3/ErbB3 protein, or Tyr1284 of human ErbB4 protein.

The epidermal growth factor (EGF) receptor is a transmembrane tyrosine kinase that belongs to the HER/ErbB protein family. Ligand binding results in receptor dimerization, autophosphorylation, activation of downstream signaling, internalization, and lysosomal degradation (1,2). The GRB2 adaptor protein binds activated EGFR at phospho-Tyr1068 (3).

The ErbB2 (HER2) proto-oncogene encodes a 185 kDa transmembrane, receptor-like glycoprotein with intrinsic tyrosine kinase activity (4). While ErbB2 lacks an identified ligand, ErbB2 kinase activity can be activated in the absence of a ligand when overexpressed and through heteromeric associations with other ErbB family members (5). Amplification of the ErbB2 gene and overexpression of its product are detected in almost 40% of human breast cancers (6). The major autophosphorylation sites in ErbB2 are Tyr1248 and Tyr1221/1222; phosphorylation of these sites couples ErbB2 to the Ras-Raf-MAP kinase signal transduction pathway (4,7).

HER3/ErbB3 is a member of the ErbB receptor protein tyrosine kinase family, but lacks tyrosine kinase activity. Tyrosine phosphorylation of ErbB3 depends on its association with other ErbB tyrosine kinases. Upon ligand binding, heterodimers form between ErbB3 and other ErbB proteins, and ErbB3 is phosphorylated on tyrosine residues by the activated ErbB kinase (8,9). There are at least 9 potential tyrosine phosphorylation sites in the carboxy-terminal tail of ErbB3. These sites serve as consensus binding sites for signal transducing proteins, including Src family members, GRB2, and the p85 subunit of PI3 kinase, which mediate ErbB-downstream signaling (10). Both Tyr1222 and Tyr1289 of ErbB3 reside within a YXXM motif and participate in signaling to PI3 kinase (11).

HER4/ErbB4, like other family members, has four ectodomains, a single transmembrane domain, and a cytoplasmic tail containing the active tyrosine kinase domain (12). By binding to neuregulins and/or EGF family ligands, ErbB4 forms either a homodimer or heterodimer with other ErbB family members, which results in receptor activation and signaling (12). ErbB4 is ubiquitously expressed with the highest expression occurring in the brain and heart. The expression of ErbB4 in breast cancer, pediatric brain cancer, and other types of carcinomas has been reported, suggesting that ErbB4 expression is involved in both normal tissue development and carcinogenesis (12).

  1. Hackel, P.O. et al. (1999) Curr Opin Cell Biol 11, 184-9.
  2. Muthuswamy, S.K. et al. (1999) Mol Cell Biol 19, 6845-57.
  3. Yarden, Y. and Sliwkowski, M.X. (2001) Nat Rev Mol Cell Biol 2, 127-37.
  4. Zwick, E. et al. (1999) Trends Pharmacol Sci 20, 408-12.
  5. Qian, X. et al. (1994) Proc Natl Acad Sci USA 91, 1500-4.
  6. Guy, P.M. et al. (1994) Proc Natl Acad Sci U S A 91, 8132-6.
  7. Dittadi, R. and Gion, M. (2000) J Natl Cancer Inst 92, 1443-4.
  8. Songyang, Z. et al. (1993) Cell 72, 767-78.
  9. Kim, H.H. et al. (1994) J Biol Chem 269, 24747-55.
  10. Kwon, Y.K. et al. (1997) J Neurosci 17, 8293-9.
  11. Rojas, M. et al. (1996) J Biol Chem 271, 27456-61.
  12. Carpenter, G. (2003) Exp Cell Res 284, 66-77.
Entrez-Gene Id
2065
Swiss-Prot Acc.
P21860
For Research Use Only. Not For Use In Diagnostic Procedures.

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