Immunoprecipitation of Spot-tagged protein.

Spot-Trap Magnetic Agarose for immunoprecipitation of Spot-tagged proteins: Coomassie blue stained gel and Western blot I: Input, FT: Flow-Through, B: Bound.

Application of Spot-tag related products.

Western blot analysis of N- and C-terminal Spot-tagged GFP added to HEK- 293T cell lysate. Detection with Spot-tag® antibody [28A5] (28a5, ChromoTek) 1:5,000 and Nano-Secondary® alpaca anti-mouse IgG1, recombinant VHH, Alexa Fluor® 488 [CTK0103, CTK0104] (sms1AF488-1, ChromoTek) 1:5,000. Western blot membrane was incubated simultaneously with the primary antibody and Nano-Secondary® (one-step staining).

Western blot analysis of C-terminal Spot-tagged GFP added to HEK-293T cell lysate. Detection with Spot-tag® antibody [28A5] (28a5, ChromoTek) 1:5,000 and anti-mouse secondary antibody HRP 1:1,000.

Western blot analysis of different cell lysates +/- 50 ng C-terminal Spot-tagged GFP: human (HEK-293T), yeast (S.cerevisiae), insect (High Five™), bacteria (E.coli) and plant (A.thaliana). Detection with Spot-tag® antibody [28A5] (28a5, ChromoTek) 1:5,000 and Nano-Secondary® alpaca anti-mouse IgG1, recombinant VHH, Alexa Fluor® 488 [CTK0103, CTK0104] (sms1AF488-1, ChromoTek) 1:5,000. Western blot membrane was incubated simultaneously with the primary antibody and Nano-Secondary® (one-step staining).

ELISA detection of N- and C-terminal Spot-tagged GFP with Spot-tag® antibody [28A5] (28a5, ChromoTek). N- and C-terminal Spot-tagged GFP was coated on a microtiter plate, blocked and detected with 1 nM Spot-tag® antibody [28A5] (28a5, ChromoTek) and anti-mouse IgG antibody AP.

Immunoprecipitation of Spot-tagged protein.

Spot-Trap Magnetic Agarose for immunoprecipitation of Spot-tagged proteins: Coomassie blue stained gel and Western blot I: Input, FT: Flow-Through, B: Bound.

Application of Spot-tag related products.

Western blot with Spot VHH.

Immunoprecipitation of Spot-tagged protein.

Immunoprecipitation of Spot-tagged protein. Input (I), non-bound (FT) and bound (B), Coomassie staining and Western blotting. Western blot indicates high effectivity.

Elution of Spot-tag fusion protein. Input (I), non-bound (FT) and eluted (E1, E2 and E3) fractions. Elutions with Spot-peptide.

Application of Spot-tag related products.

STED: IF of Spot-tagged Actin-Chromobody with Spot-Label Atto594 bivalent (1:1,000). Gated STED images were acquired with a Leica TCS SP8 STED 3X microscope with pulsed White Light Laser excitation at 590 nm and pulsed depletion with a 775 nm laser. Objective: 100x Oil STED White, NA: 1.4. Pixel size: 21 x 21 nm; z-Step size of z-Stacks: 0.16 μm. Images were deconvolved with Huygens Professional (SVI). STED images were recorded at the Core Facility Bioimaging at the Biomedical Center, LMU Munich.

Application of Spot-tag related products.

Widefield images of HeLa cells expressing Tom70-EGFP-Spot-Tag immunostained with Spot-Label ATTO594, bivalent (1:5,000). Left: GFP channel, middle: channel for ATTO594, right: overlay, also with DAPI. Scale bar, 10 μm.

Confocal images of HeLa cells expressing Tom70-EGFP-Spot-Tag immunostained with Spot-Label ATTO594 bivalent (1:1,000). Left: EGFP fluorescence, middle: ATTO594 signal, right: merge. Scale bar, 10 μm. The images were acquired with Leica TCS SP8 microscope, 100X oil objective, at the Core Facility Bioimaging at the Biomedical Center, LMU Munich, and deconvolved with Huygens Professional (SVI). Left: GFP fluorescence, middle: ATTO594 signal, right: merge. Scale bar, 10 μm.

SPOT tag related products

Left: HeLa cells transiently expressing pSpot Spot-Tag?-Actin (ev-31) were immunostained with Spot-Label? Alexa Fluor? 568 (red, ebAF568, 1:800) and DAPI (blue). Right: Control staining with Spot-Label? Alexa Fluor? 568 (red, ebAF568, 1:800) and DAPI (blue) of untransfected HeLa cells showing no unspecific background. Scale bar, 10 μm. Images were acquired with the Thermo Scientific CellInsight CX7, 20X objective.

Make the most of your time & gain about 40 hours with the iST_Spot-Trap Kit.

Schematic workflow of iST_Spot-Tap_kit.

Preomics Technology Priciple.