At D11 or D21, cells were plated onto Greiner Bio-One Cellstar 96-well μclear Flat Bottomed Microplates (Fisher Scientific 07-000-166) for immunocytochemistry at a density of 90,000 cells/cm2. Cells were fixed 24 hours later with 4% paraformaldehyde for 30 minutes at room temperature. Cells were permeabilized in PBS with 0.2% Triton X-100 (PBS+) for 20 minutes and blocked in PBS with 5% donkey serum and 0.05% TritonX-100 (PBS++) for 2 hours. Primary antibodies were diluted in PBS++ and incubated on cells overnight at 4°C. Primary antibodies included: gt anti-SOX1 (R&D Systems AF3369) diluted 1:100, rb anti-PAX6 (BioLegend 901301) diluted 1:200, rb anti-Ki67 (Abcam Ab16667) diluted 1:100, rabbit anti-FOXG1 (Abcam AB18259) diluted 1:100, and mouse anti-MAP2 (ThermoFisher Scientific 131500) diluted 1:500. Secondaries used included: Alexa Fluor 488 AffiniPure donkey anti-goat IgG, donkey anti-mouse IgG (Jackson ImmunoResearch Laboratories 715-545-150), and donkey anti-rabbit IgG (Jackson ImmunoResearch Laboratories 715-545-152). Secondaries were diluted 1:800 in PBS++ and incubated for 3 hours at room temperature. Cells were incubated for 5 minutes with Hoechst nuclear dye (ThermoFisher Scientific H3570) at 2μg/mL and stored in PBS, protected from light, at 4°C until imaging.
Mouse anti map2
Manufactured by Thermo Fisher Scientific
Sourced in United States
Mouse anti-MAP2 is a primary antibody used for the detection of Microtubule-Associated Protein 2 (MAP2) in various sample types. MAP2 is a structural protein found in neurons and plays a role in the stabilization and organization of microtubules. The mouse anti-MAP2 antibody can be used in techniques such as immunohistochemistry, immunocytochemistry, and Western blotting to identify and localize MAP2 in research applications.
Automatically generated - may contain errors
Lab products found in correlation
Lsm 780 confocal microscope, by Zeiss (3 mentions)
Hoechst 33342, by Thermo Fisher Scientific (3 mentions)
4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (2 mentions)
Ab18259, by Abcam (1 mentions)
Gt anti sox1, by R&D Systems (1 mentions)
Rb anti ki67, by Abcam (1 mentions)
Hoechst nuclear dye, by Thermo Fisher Scientific (1 mentions)
Donkey anti rabbit igg, by Jackson ImmunoResearch (1 mentions)
Rabbit anti dcx, by Cell Signaling Technology (1 mentions)
Alexa fluor 488 conjugated goat anti rabbit igg, by Thermo Fisher Scientific (1 mentions)
Alexa fluor 555 conjugated goat anti mouse igg, by Thermo Fisher Scientific (1 mentions)
Paraformaldehyde (pfa), by Thermo Fisher Scientific (1 mentions)
Igepal ca 630, by Merck Group (1 mentions)
Chicken anti map2, by Abcam (1 mentions)
Alexa fluor 488 goat anti mouse, by Thermo Fisher Scientific (1 mentions)
Alexa fluor 488 goat anti chicken, by Thermo Fisher Scientific (1 mentions)
Rabbit anti glutamate, by Merck Group (1 mentions)
Rabbit anti vglut1, by Synaptic Systems (1 mentions)
Hoechst 33258, by Merck Group (1 mentions)
Rabbit anti pax6, by Fortrea (1 mentions)
6 protocols using mouse anti map2
1
Quantifying Neural Progenitor Markers
2
Immunostaining of Neuronal Markers in Mouse Brain
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
After the VLIUS stimulation, the mice were sacrificed and perfused with 10 % formaldehyde/PBS. The brain was then harvested and fixed with 10% formaldehyde/PBS at room temperature. Samples were embedded in paraffin and serial 7 μm transverse sections were mounted on slides. The samples were deparaffinized, rehydrated, antigen retrieved (100℃, 20 min) and washed in PBST. Slices were blocked with 10% newborn calf serum (NCS) and 1% BSA in PBST for 1 hr, incubated with primary antibody overnight at 4℃. After washing with PBST, the samples were incubated with the secondary antibody for 1 hr at room temperature, washed with PBST and mounted with EverBrite Hardset Mounting Medium containing DAPI to label the nuclei (Biotium). Slides were viewed, and images were captured with LSM780 confocal microscope (Zeiss, Jena, Germany). The primary antibodies used for immunostaining and their dilutions were as follows: rabbit anti-DCX (1:200, Cell signaling), mouse anti-MAP2 (1:200, Thermo). The secondary antibodies used were Alexa Fluor 488-conjugated goat anti-rabbit IgG (1:100, Thermo) and Alexa Fluor 555-conjugated goat anti-mouse IgG (1:100, Thermo).
3
Characterization of iPSC-Derived Neurons
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
To characterize the iPSCs-derived neurons, cells were fixed using 4% paraformaldehyde (ThermoFisher Scientific, 28908, Waltham MA, USA) and permeabilized using 0.2% Igepal CA-630 (Sigma-Aldrich, I3021, Darmstadt, Germany). Neurons were stained with mouse anti-MAP2 (ThermoFisher Scientific, MA5-12826, Waltham MA, USA) and goat anti-mouse Alexa Fluor 488 (ThermoFisher Scientific, A-11001, Waltham MA, USA), or chicken anti-MAP2 (Abcam, ab92434, Cambridge, United Kingdom) and goat anti-chicken Alexa Fluor 488 (ThermoFisher Scientific, A-11039, Waltham MA, USA) to depict their cytoskeletal structure. Nuclei were visualized using the nuclear counterstain DAPI (ThermoFisher Scientific, D1306, Waltham MA, USA). Glial progenitors that were generated from the NPCs were also resolved with mouse anti-MAP2 and goat anti-mouse Alexa Fluor 488 in culture24 (link).
4
Immunofluorescence Analysis of Neural Cell Markers
5
Immunofluorescence Labeling Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Immunofluorescence procedures were performed as previously reported31 (link). Coverslips with cells or tissue sections were labeled with primary antibodies overnight at 4 °C. Antibodies used included rabbit anti-MAG (Thermo Fisher Scientific), mouse anti-Tubulin III (R&D Systems), rabbit anti-Tubulin III (Abcam, Cambridge, MA, USA), mouse anti-GFAP (Millipore, Billerica, MA, USA), rabbit anti-p-PKCγ (Abcam), mouse anti-MAP2 (Thermo Fisher Scientific), mouse anti-β-catenin (Abmart, Shanghai, China), mouse anti-Nestin (Abcam), rabbit anti-Rip (a gift from Dr. Scott R. Whittemore, University of Louisville) and mouse anti-hemagglutinin (HA)-tag (Abmart). Cell nuclei were counterstained with Hoechst 33342 (Invitrogen). The staining was visualized using an EVOS FL microscope (Life technology, Gaithersburg, MD, USA) or a laser scanning confocal microscope (Leica Microsystems GmbH, Mannheim, Germany).
6
Immunostaining of Astrocytes, Neurons, and Nuclei
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Samples were immunostained following the protocol reported in Girardin et al. (2022 (link)). After 4% paraformaldehyde (1.00496, Sigma Aldrich) fixation, 0.2% Triton X-100 (X100, Sigma Aldrich) permeabilization and blocking of non-specific binding with 0.2% Triton-X and 3% bovine serum albumin (BSA, A7906, Sigma Aldrich), samples were incubated overnight at 4 °C with a solution of PBS containing 0.2% Triton-X, 3% BSA and the primary antibodies. Primary antibodies were chicken anti-GFAP (1:1,000, AB4674, Abcam), rabbit anti-S100β (AB34686, Abcam), and mouse anti-MAP2 (1:250, 13–1,500, Thermo Fisher). On the next day, samples were rinsed with PBS and incubated with a secondary antibody solution in PBS for 2 h at room temperature. Secondary antibodies were goat anti-mouse DyLight 405 (1:800, 35500BID, Thermo Fisher), goat anti-chicken Alexa Fluor 488 (1:800, A11039, Thermo Fisher) and donkey anti-rabbit Alex Fluor Plus 647 (1:800, A32795, Thermo Fisher). Depending on the experiment, Hoechst 33342 (H3570, Thermo Fisher) was added to the antibody solution at a working concentration of 2 μM instead of the DyLight 405 antibody. The samples were rinsed three times with PBS and left in PBS for imaging.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!