Anti-mouse IgG1 Nano-Secondary: Well-defined and characterized immunostaining. Primary anti-mouse IgG1 antibody (grey) with 2X2 monoclonal mouse Fc- specific Nanobodies (green) bound. In total, 8 fluorophores (red stars) label the mouse IgG1 primary antibody.
Immunostaining with Alpaca anti-mouse IgG1 Alexa Fluor 647 Nano-Secondaries for FACS. Jurkat CD3+ and Jurkat CD3- cell lines were mixed and immunostained live with anti-CD3 mouse IgG1 + alpaca anti-mouse IgG1 VHH Alexa Fluor 647 (1:600). CD3- cells were pre-stained with Calcein. Two cell populations can be clearly distinguished on the dot-plot: Alexa Fluor 647-positive Calcein-negative cells and Alexa Fluor 647-negative Calcein-positive cells.
Multiplexed immunostaining of HeLa cells with 3 subclass-specific alpaca anti-mouse Nano-Secondaries. Grey: Mouse IgG1 anti-Vimentin + alpaca anti-mouse IgG1 VHH Alexa Fluor? 647. Green: Mouse IgG2b anti-Lamin + alpaca anti-mouse IgG2b VHH Alexa Fluor? 488. Red: Mouse IgG3 anti-MOT + alpaca anti-mouse IgG3 VHH Alexa Fluor? 568. Scale bar, 10 μm. Images were recorded at the Core Facility Bioimaging at the Biomedical Center, LMU Munich.
Western blot analysis of EGFP (EGFP-250, ChromoTek) added to HEK293T cell lysate. Detection with anti-GFP mouse IgG 1 antibody and alpaca anti-mouse IgG1 VHH Alexa Fluor? 647.
One-step staining (left) vs. sequential staining (right) of HeLa cells with anti-COX4 (mitochondria) mouse IgG1 monoclonal primary antibody + alpaca anti-mouse IgG1 VHH Alexa Fluor? 647 (magenta). Cell nuclei are stained with DAPI (blue). Scale bar, 20 μm.
The anti-mouse IgG1 Nano-Secondary is subclass-specific and does not cross-react with IgGs from other commonly used species (here rabbit) and with mouse IgG2b and IgG3 subclasses.
Multiplexed immunostaining of HeLa cells with 3 subclass-specific alpaca anti-mouse Nano-Secondaries. Grey: Mouse IgG1 anti-Vimentin + alpaca anti-mouse IgG1 VHH Alexa Fluor? 647. Green: Mouse IgG2b anti-Lamin + alpaca anti-mouse IgG2b VHH Alexa Fluor? 488. Red: Mouse IgG3 anti-MOT + alpaca anti-mouse IgG3 VHH Alexa Fluor? 568. Scale bar, 10 μm. Images were recorded at the Core Facility Bioimaging at the Biomedical Center, LMU Munich.
HeLa cells were immunostained with anti-Vimentin mouse IgG1 antibody and alpaca anti-mouse IgG1 VHH Alexa Fluor? 647 (grey). Nuclei were stained with DAPI, blue. Scale bar, 10 μm. Images were recorded at the Core Facility Bioimaging at the Biomedical Center, LMU Munich.
One-step immunostaining is the simultaneous incubation of mouse IgG1 primary antibody and anti-mouse IgG1 Nano-Secondary. This method reduces incubation and hands-on time. Simultaneous incubation also supports multiplexing, tissue penetration, and cell staining for flow cytometry.
Flow cytometry with anti-mouse Nano-Secondaries. Jurkat CD3+ and Jurkat CD3- cell lines were mixed and immunostained live with anti-CD3 mouse IgG1 and alpaca anti-mouse IgG1 VHH Alexa Fluor? 647 (1:600). Two cell populations can be clearly distinguished with a 2-log shift: CD3-positive cells (in green) and CD3-negative cells (in grey).
