41-2100

antibody from Invitrogen Antibodies

Targeting: MAPRE1 EB1

Provider product page for 41-2100

Western blot

ELISA

Other assay

Antibody data

Antibody Data
Antigen structure
References [2]
Comments [0]
Validations
Western blot [2]
Other assay [2]

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Product number: 41-2100 - Provider product page
Provider: Invitrogen Antibodies
Product name: EB1 Monoclonal Antibody (1A11/4)
Antibody type: Monoclonal
Antigen: Recombinant full-length protein
Reactivity: Human, Mouse, Rat, Canine
Host: Mouse
Isotype: IgG
Antibody clone number: 1A11/4
Vial size: 100 µg
Concentration: 0.5 mg/mL
Storage: -20°C

MAPRE1 protein structure - 41-2100 shown in red.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis was performed on membrane enriched extract (30 µg lysate) of HCT 116 (Lane 1), HeLa (Lane 2), U-87 MG (Lane 3), SW480 (Lane 4), HT-29 (Lane 5), COLO 205 (Lane 6) and MKN45 (Lane 7).The blot was probed with EB1 Mouse monoclonal Antibody (Product # 41-2100, 2 µg/mL) and detected by chemiluminescence using Goat anti-Mouse IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A28177, 0.25 µg/mL, 1:4000 dilution). A 32 kDa band corresponding to EB1 was observed in cell lines tested. Known quantity of protein samples were electrophoresed using Novex® NuPAGE® 4-12 % Bis-Tris gel (Product # NP0322BOX), XCell SureLock™ Electrophoresis System (Product # EI0002) and Novex® Sharp Pre-Stained Protein Standard (Product # LC5800). Resolved proteins were then transferred onto a nitrocellulose membrane with iBlot® 2 Dry Blotting System (Product # IB21001). The membrane was probed with the relevant primary and secondary Antibody following blocking with 5% skimmed milk. Chemiluminescent detection was performed using Pierce™ ECL Western Blotting Substrate (Product # 32106).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Knockdown of EB1 was achieved by transfecting HeLa cells with EB1 specific validated siRNAs (Silencer® select Product # s22674, s22675). Western blot analysis (Fig 1) was performed using membrane enriched extracts from the EB1 knockdown cells (lane 3), non-specific scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blots were probed with EB1 Monoclonal Antibody (Product # 41-2100, 1 µg/mL) and Goat anti-Mouse IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A28177, 0.25µg/mL, 1:4000 dilution). Densitometric analysis of this western blot is shown in histogram (Fig 2). Decrease in signal upon siRNA mediated knock down confirms that antibody is specific to EB1 .

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 1. Stable MTs Mediate EB-Independent HSV-1 Infection of SK-N-SHs (A) NHDFs or SK-N-SHs treated with non-targeting (ctrl), EB1, or CLIP-170 (CLIP) siRNAs were mock-infected or infected with HSV-1 at MOI 10 for 5 h and analyzed by WB. (B) SK-N-SHs treated with the indicated siRNAs were infected, as in (A). (C) SK-N-SHs treated with independent EB2 siRNAs (I or II) were infected, as in (A). (D) SK-N-SHs were treated with 100 nM BafA or DMSO and infected at MOI 10 with VSV for 4 h or HSV-1 for 5 h. (E) NHDFs or SK-N-SHs analyzed by WB using the indicated antibodies. (F) NHDFs or SK-N-SHs stained for tyrosinated (Tyr), detyrosinated (Detyr), and acetylated (Ac) tubulin. Nuclei were stained with Hoechst. (G) NHDFs or SK-N-SHs treated with 500 nM DMSO or 10 muM nocodazole were infected at MOI 20 with HSV-1 for 4 h in the presence of 1 mug/mL actinomycin D. Fixed cells were stained for VP5 and with Hoechst. Assessed for the accumulation of VP5 over 2 biological replicates were >= 165 NHDF or >= 190 SK-N-SH nuclei; error bars, SEMs; *p < 0.05, **p < 0.01, N.S., not significant; unpaired 2-tailed t test. (H) Cells treated as in (G) were infected at MOI 10 with HSV-1 for 5 h. All of the experiments represent >=3 replicates unless indicated. See also Figures S1 and S2 .

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 3. TACC3 Regulates MT Dynamics and chTOG Localization (A) SK-N-SHs were treated with siRNAs and then fixed and stained for tyrosinated (Tyr) and detyrosinated (Detyr) tubulin, EB1, and Hoechst. Representative cells are shown. (B) Average number of EB1 comets per cell treated as in (A). Total number of EB1 comets in 30 cells per siRNA over 3 biological replicates; error bars, SEMs; **p < 0.01; unpaired 2-tailed t test. (C) SK-N-SHs were transduced with pQC, pQC-TACC3, or pQCXIP-FLAG-TACC3 (pQC-FLAG-TACC3) retroviral vectors and analyzed by WB. (D) SK-N-SHs transfected with pQC-FLAGTACC3 and fixed and stained for FLAG and with Hoechst. (E) SK-N-SHs transduced as in (C) were fixed and stained for TACC3 and Hoechst. (F) SK-N-SHs were transfected with pQC or pQC-FLAG-TACC3, fixed after 3 days, and stained for FLAG (red), EB1 (green), and with Hoechst. Enlarged image shows elongated EB1 staining patterns, indicative of reduced MT polymerization, in cells expressing FLAG-TACC3. (G) SK-N-SHs as in (F) were fixed and stained for FLAG (green) and chTOG (red) and with Hoechst. (H) SK-N-SHs treated with control or TACC3 siRNA and analyzed by WB. (I) siRNA-treated SK-N-SHs fixed and stained for chTOG (red) and with Hoechst. (J) Average corrected total fluorescence intensity of nuclear chTOG in SK-N-SHs treated with siRNAs; >= 255 cells per siRNA over 3 biological replicates; error bars, SEMs; **p < 0.01; unpaired 2-tailed t test. (K) SK-N-SHs were treated with siRNAs, fixed and sta