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Rabbit anti foxp1

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Rabbit anti-FoxP1 is an antibody that recognizes the FoxP1 protein. FoxP1 is a transcription factor that plays a role in the regulation of gene expression.

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Lab products found in correlation

Rabbit anti olig2, by Merck Group (2 mentions) Advantage cdna pcr kit, by Takara Bio (1 mentions) Mouse anti gfp, by Merck Group (1 mentions) Rabbit anti krox20, by Fortrea (1 mentions) Rabbit anti βiii tubulin, by Fortrea (1 mentions) Alexa fluor 488 conjugated donkey anti rabbit, by Jackson ImmunoResearch (1 mentions) Triton x 100, by Merck Group (1 mentions) Goat anti ephb2, by R&D Systems (1 mentions) Rabbit anti ph3, by Merck Group (1 mentions) Rabbit anti epha4, by Santa Cruz Biotechnology (1 mentions) Lipofectamine 3000 reagent, by Thermo Fisher Scientific (1 mentions) Chicken anti gfp antibody, by Abcam (1 mentions) 4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (1 mentions) Rabbit anti aldh1l1, by Abcam (1 mentions) Leica dm5500 b epifluorescence microscope, by Leica Microsystems (1 mentions) Mouse anti gad67, by Merck Group (1 mentions) Chicken anti map2, by Abcam (1 mentions) Mouse anti tuj1, by Abcam (1 mentions) Dylight 488 or 549, by Jackson ImmunoResearch (1 mentions) C9100 13 camera, by Hamamatsu Photonics (1 mentions)

8 protocols using rabbit anti foxp1

1

Immunohistochemical and in situ Hybridization Protocols

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Embryos were obtained and processed for immunohistochemistry or in situ hybridization as described previously (Song, et al., 2009 (link)). The following antibodies were used: rabbit and guinea pig anti-Hb9 (Thaler, et al., 1999 (link)), guinea pig anti-Lhx3 (Sharma, et al., 1998 (link)), guinea pig anti-Chx10 (Thaler, et al., 2002 (link)), rabbit anti-Foxp1 (Abcam), rabbit anti-Krox20 (Covance), rabbit and guinea pig anti-Isl1/2 (Ericson, et al., 1992 (link)), mouse anti-Neurofilament (DSHB), mouse anti-GFP (Sigma), rabbit anti-β-III-tubulin (Covance), rabbit anti-β-galactosidase (Cappel) antibodies. For immunocytochemistry, dissociated cultured cells were fixed and immunostained with antibodies including rabbit anti-Robo1 and Robo2 (kind gift of Dr. Elke Stein, Yale) and mouse anti-Isl1 (DSHB). Previous characterization of the Robo1 and Robo2 antisera confirmed that specific labeling was lost in homozygous mutants. For in situ hybridization, embryonic cDNA at E10.5 or E12.5 was used to generate riboprobes using an Advantage cDNA PCR kit (Clonetech).
Lee H., Kim M., Kim N., Macfarlan T., Pfaff S.L., Mastick G.S, & Song M.R. (2015). Slit and semaphorin signaling governed by Islet transcription factors positions motor neuron somata within the neural tube. Experimental neurology, 269, 17-27.
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2

Retrograde Viral Tracing of RMTg Inputs

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Subjects were deeply anesthetized by inhaled isoflurane and transcardially perfused using 0.9% saline followed by 10% formalin. Brains were removed and stored in 10% formalin overnight before being transferred to 20% sucrose with 0.05% sodium azide for cryoprotection. Tissue was collected in 40μm-thick sections on a cryostat or freezing microtome, and floating sections were processed using immunohistochemistry to verify virus expression in the RMTg and accurate placement of cannulae or optical fibers in the VTA. Tissue was incubated overnight in PBS with 0.25% Triton X-100 (Sigma-Aldrich) and primary antibody for rabbit anti-GFP (1:50K, Abcam), mouse anti-tyrosine hydroxylase (TH; 1:10K, Millipore Inc.), or rabbit anti-FoxP1 (1:20K, Abcam). Fluorescence was visualized using 30-min secondary incubation in either Alexa Fluor 488-conjugated donkey anti-rabbit or Cy3-conjugated donkey anti-mouse secondary (1:1000, Jackson Immunoresearch). Following each incubation step, tissue was rinsed 3X in PBS at 1min/wash. Data were included from animals showing GFP+ cells clustered bilaterally in the region corresponding to the FoxP1-positive RMTg [15 (link)], with dense labelling of axon terminals in the TH-positive VTA.
Vento P.J., Watson J.R., Pullmann D., Black S.L., Tomberlin J.S, & Jhou T.C. (2023). Pumping the brakes: rostromedial tegmental inhibition of compulsive cocaine seeking. bioRxiv.
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3

Molecular mechanisms of Efnb2 and Efnb3 in mouse embryonic development

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All analyses for Efnb2 cKO were performed on control and mutant littermates collected from at least two different litters. On the other hand, control and Efnb3 mutant embryos were collected from independent litters. The number of embryo analyzed for each immunostaining and each developmental stage is indicated in the figure legends. To avoid bias in rostro-caudal axis, data was collected on thick vibratome sections covering the entire brachial region (600 μm). Antibody staining was performed following standard protocol on 70μm vibratome sections of mouse embryos at brachial level. For BrdU incorporation, pregnant dams were injected with BrdU (10mg/ml; 100mg/kg) with intraperitoneal injection. After 1 h, embryos were dissected in cold PBS and processed for subsequent immunostaining.
Antibodies used were: goat anti-Nkx2.2 (1/100, Santa Cruz Biotechnology); rabbit anti-Olig2 (1/1000, Sigma); mouse anti-Islet1/2, 39-4D5 (1/50, DSHB); rabbit anti-Foxp1 (1/200, Abcam), rabbit anti-P-H3 (1/1000, Millipore), rabbit anti-EphA4 (1/100, Santa Cruz Biotechnology), goat anti-EphB2 (1/50, R&D Systems), Tuj1 (1/1000, Covance). All secondary antibodies were from Jackson ImmunoResearch (1/1000).
Laussu J., Audouard C., Kischel A., Assis-Nascimento P., Escalas N., Liebl D.J., Soula C, & Davy A. (2017). Eph/Ephrin Signaling Controls Progenitor Identities In The Ventral Spinal Cord. Neural Development, 12, 10.
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4

Visualizing FOXP1 Protein Localization

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Transient transfections were carried out as previously described46 (link). Briefly, HeLa cells were grown in 24 well plates on coverglass to 60–80% confluency. Cells were transfected with 500 ng pcDNA 3.1_FoxP1 WT, p.R525*, or p.R525Q separately using Lipofectamine 3000 reagents (Thermo Fisher Scientific) following the manufacturer’s suggested protocol. After 48 hours, cells were fixed with 3% paraformaldehyde for 20 minutes. Primary antibody incubation was performed with rabbit anti-FOXP1 (Abcam) and chicken anti-GFP antibodies (Abcam) followed by a secondary antibody incubation using anti-rabbit-568, anti-chicken-488 (Invitrogen), and DAPI (Life Technologies). Imaging for cellular localization was carried out on a Nikon Ni-E microscope equipped with a Photometrics CoolSNAP Myo camera. Exposures for each channel were 700 ms (DAPI), 900 ms (GFP), and 300 ms (Texas Red).
Johnson T.B., Mechels K., Anderson R.E., Cain J.T., Sturdevant D.A., Braddock S., Pinz H., Wilson M.A., Landsverk M., Roux K.J, & Weimer J.M. (2018). Characterization of a recurrent missense mutation in the forkhead DNA-binding domain of FOXP1. Scientific Reports, 8, 16161.
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5

Immunofluorescence Imaging of Neural Markers

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Tissue cryosections were washed in 1X PBS then blocked for 1h at room temperature (RT) in 1X PBS/0.3% Triton X-100/3% normal donkey or goat serum (blocking solution). Slides were incubated overnight at 4°C with blocking solution containing dilutions of the following antibodies: rabbit anti-ALDH1L1 (Abcam) 1:500; rabbit anti-cleaved caspase-3 (Biocare Medicare) 1:250; rabbit anti-FoxP1 (Abcam Inc.) 1:400; mouse anti-GAD67 (EMD Millipore) 1:5000; rabbit anti-glycine (Millipore) 1:100; chicken anti-MAP2 (Abcam Inc.) 1:5000; rabbit anti-Olig2 (EMD Millipore) 1:250; mouse anti-TUJ1 (Abcam) 1:500. Sections were washed in 1X PBS and incubated for 1h with secondary antibodies conjugated to DyLight 488 or 549 (Jackson Immunoresearch) at a 1:500 dilution. All slides were counterstained with 4′, 6-diamidino-2-phenylindole (DAPI) or NeuroTrace fluorescent Nissl stain (Molecular Probes). After staining, sections were mounted with ProLong Gold to preserve the fluorescent signals and imaged using a Leica DM5500B epifluorescence microscope (Leica Microsystems, Exton, PA) or an inverted Zeiss Axio Observer on a PerkinElmer UltraVIEW VoX spinning disk confocal with a Hamamatsu C9100-13 camera and Volocity software.
Altieri S.C., Zhao T., Jalabi W., Romito-DiGiacomo R.R, & Maricich S.M. (2016). En1 is necessary for survival of neurons in the ventral nuclei of the lateral lemniscus. Developmental neurobiology, 76(11), 1266-1274.
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6

Immunohistochemistry and in situ Hybridization

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Immunohistochemistry was performed as described previously [35 (link)]. Following antibodies were used: mouse anti-Isl1 (DSHB), mouse anti-Isl2 (DSHB), rabbit anti-Isl1/2 [22 (link)], mouse anti-MNR2 (DSHB), rabbit anti-GFP (Invitrogen), rabbit anti-Foxp1 (Abcam), rabbit anti-Sox1 (Cell signaling), guinea pig anti-Lhx3 [71 (link)], guinea pig anti-Chx10 [71 (link)], guinea pig anti-Olig2 [36 (link)] and mouse anti-Neurofilament (DSHB). For wholemount immunostaining, Day 4.5 chick embryos electroporated at Day 2 were fixed and incubated with primary antibodies for 3 days and secondary antibodies for 1 day. For in situ hybridization, transverse sections were hybridized with digoxigenin-labeled probes specific for mouse Phox2a (full CDS), mouse OC-1 (partial CDS, 469–1263 bp). All images were captured with epifluorescent microscope or confocal microscope (Zeiss).
Kim N., Park C., Jeong Y, & Song M.R. (2015). Functional Diversification of Motor Neuron-specific Isl1 Enhancers during Evolution. PLoS Genetics, 11(10), e1005560.
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7

Co-Immunoprecipitation of AKT-Regulated Proteins

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HEK293T cells (ATCC, #CRL-3216) were cultured in DMEM media supplemented with 10% FBS. For co-immunoprecipitation, HEK293T cells were seeded on 10 cm tissue culture dishes, cultured in DMEM media supplemented with 10% FBS, and transfected with the expression vectors tagged with Flag, and HA using Superfect (Qiagen) or calcium phosphate. Cells were treated with AKT1/2 kinase inhibitor, iAKT1/2 (Sigma Aldrich, A6730) at 10 μM for 20 hr and then cells were harvested and lysed in IP buffer (20 mM Tris-HCl, pH 8.0, 0.5 % NP-40, 1 mM EDTA, 150 mM NaCl, 2 mM PMSF, 10% Glycerol, 4 mM Na3VO4, 200 mM NaF, 20 mM Na-pyroPO4, and protease inhibitor cocktail). In these studies, immunoprecipitations were performed with mouse anti-HA antibody (Covance). Immunoblotting assays were performed using goat anti-Gli3 (R and D Systems, AF3690, 1:250), rabbit anti-ARHGAP36 (Sigma, HPA-002064, 1:2000), mouse anti-HA (Covance, 1:5000), rabbit anti-β-tubulin (Santa Cruz, sc-9104, 1:2000), rabbit anti-pSER (Cell Signaling, #9651, 1:5000), mouse anti-TuJ1 (Covance, 1:5000), rabbit anti-FoxP1 (abcam, ab16645, 1:1000) and anti-pCREB (Cell Signaling, 9198S, 1:1000) antibodies.
Nam H., Jeon S., An H., Yoo J., Lee H.J., Lee S.K, & Lee S. (2019). Critical roles of ARHGAP36 as a signal transduction mediator of Shh pathway in lateral motor columnar specification. eLife, 8, e46683.
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8

Multimarker Fluorescence Immunocytochemistry

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Fluorescent immunocytochemistry followed standard protocols with primary antibodies for rabbit anti-FoxP1 (1:500; Abcam), mouse anti-FoxP1 (JC12) (1:500; Abcam), mouse anti-β-III-tubulin (1:1000; Sigma), mouse anti-Map2ab (1:500; Sigma), mouse anti-DARPP-32 (1:20,000; a gift from Prof H. Hemmings), mouse anti-GFAP (1:500; Abcam), rabbit anti-GAD65/67 (1:2000; Millipore), rabbit anti-met-enkephalin (1:15,000; Millipore), rat anti-CTIP2 (1:500, Abcam) and rat anti-BrdU (1:200, Oxford Bio). For double labelling, the two primary antibodies that had been raised in different species were added at the same time. Appropriate fluorescent-labelled secondary antibodies (Life technologies) were applied, followed by the nuclear stain Hoechst. Fluorescent staining was visualised using a Leica DRMBE microscope at 560 nm (red), 494 nm (green) and 346 nm (blue). Cell counts were undertaken at 40 × magnification using a counting grid. For unbiased sampling, 5 fields were chosen at random from which to take counts. Pseudocolour fluorescent images were obtained using Openlab 2.1 image analysis software and colour images were processed using Adobe Photoshop.
Precious S.V., Kelly C.M., Reddington A.E., Vinh N.N., Stickland R.C., Pekarik V., Scherf C., Jeyasingham R., Glasbey J., Holeiter M., Jones L., Taylor M.V, & Rosser A.E. (2016). FoxP1 marks medium spiny neurons from precursors to maturity and is required for their differentiation. Experimental Neurology, 282, 9-18.
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