We collected the culture progeny of MuSCs from aged Myf5nLacZ/+/Luciferase double-transgenic mice23 (link),43 (link) by incubation with 0.1% trypsin in PBS for 2 min at 37 °C and transplanted them into tibialis anterior muscles of hindlimb-irradiated NOD/SCID mice. One month after transplant, we injected notexin to damage recipient muscles and activate MuSCs in vivo. Four days later, we collected, fixed, and cryosectioned recipient muscles, as described above. We performed immunohistological analysis of transverse tissue sections to detect β-galactosidase+ cells (indicating a donor-derived cell expressing Myf5, a marker of MuSC activation) in the satellite cell position within the myofiber basal lamina, as defined by laminin staining. We stained sections with anti-Laminin (Millipore, clone A5, catalog # 05-206, 1:250) and anti-β-galactosidase (Invitrogen, catalog # A11132, 1:100) primary antibodies and then with appropriate secondary antibodies (Invitrogen). We counter-stained nuclei with Hoechst 33342 (Invitrogen). We acquired images with an AxioPlan2 epi-fluorescent microscope (Carl Zeiss) with Plan NeoFluar 10×/0.30NA or 20×/0.75NA objectives (Carl Zeiss) and an ORCA-ER digital camera (Hamamatsu). We captured digital images in OpenLab software (Improvision) and assembled them using Photoshop software (Adobe) with consistent contrast adjustments across all images.
Anti laminin
Manufactured by Merck Group
Sourced in United States, United Kingdom, Denmark
Anti-laminin is a laboratory reagent used to detect and quantify the presence of laminin, a protein found in the extracellular matrix. It is commonly used in research applications to study cell-matrix interactions, tissue development, and various pathological conditions.
Automatically generated - may contain errors
Lab products found in correlation
Anti fibronectin, by Merck Group (8 mentions)
4 6 diamidino 2 phenylindole (dapi), by Merck Group (6 mentions)
Alexa fluor 488, by Thermo Fisher Scientific (6 mentions)
Orca er digital camera, by Hamamatsu Photonics (5 mentions)
Anti gfp, by Thermo Fisher Scientific (5 mentions)
4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (5 mentions)
Anti pax7, by Developmental Studies Hybridoma Bank (5 mentions)
Openlab software, by PerkinElmer (4 mentions)
Hoechst 33342, by Thermo Fisher Scientific (4 mentions)
Anti collagen 1, by Abcam (4 mentions)
Anti collagen 4, by Abcam (4 mentions)
Rhodamine phalloidin, by Thermo Fisher Scientific (4 mentions)
Axioplan2 epifluorescent microscope, by Zeiss (3 mentions)
Anti myod, by Santa Cruz Biotechnology (3 mentions)
Anti ki67, by Abcam (3 mentions)
Native laminin, by Merck Group (3 mentions)
Vectashield with dapi, by Vector Laboratories (3 mentions)
Alexa fluor conjugated secondary antibody, by Thermo Fisher Scientific (3 mentions)
Appropriate secondary antibody, by Thermo Fisher Scientific (2 mentions)
Anti β galactosidase, by Thermo Fisher Scientific (2 mentions)
105 protocols using anti laminin
1
Quantifying Activated MuSCs in Transplanted Muscle
2
Immunohistochemical Analysis of Donor-Cell Transgene
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
We collected and prepared recipient tibialis anterior muscle tissues for histology to analyze the expression of donor-cell transgene products as previously described23 (link). We incubated transverse sections with anti-Laminin (Millipore, clone A5, catalog # 05-206, 1:250), anti-GFP (Invitrogen, catalog # A11122, 1:200), and/or anti-Luciferase (Abcam, catalog # ab81822, 1:100) primary antibodies and then with appropriate secondary antibodies (Invitrogen). We counter-stained nuclei with Hoechst 33342 (Invitrogen) or Topro3 (Invitrogen). We acquired images with an AxioPlan2 epi-fluorescent microscope (Carl Zeiss Microimaging, Thornwood, NY) with Plan NeoFluar 10x/0.30NA or 20x/0.75NA objectives (Carl Zeiss) and an ORCA-ER digital camera (Hamamatsu Photonics). We captured digital images in OpenLab software (Improvision) and assembled multi-panel figures using Photoshop software (Adobe) with consistent contrast adjustments across all images from the same stain.
3
Glioblastoma Xenograft Model in SCID Mice
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
All animal protocols were approved by the Johns Hopkins School of Medicine Animal Care and Use Committee. Each 6-8 week old female BALB/c strain immunodeficient (SCID) mouse received 10,000 viable neurosphere cells in 2 μL PBS by stereotactic injection to the right caudate/putamen (AP = 0 mm, ML = −2.5 mm, DV = −3.0 mm). After 7 weeks, mice were sacrificed and perfused with 4% paraformaldehyde; the brains were removed for histological analysis. Tumor sizes were quantified by measuring maximum tumor volume on hematoxylin and eosin–stained brain coronal sections using computer-assisted morphometry (MCID software) and then applying the formula Volume = (square root of maximum cross-sectional area).3 ,20 (link) The primary antibodies used for immunofluorescent staining are the following: monoclonal anti-Ki67 (BD Biosciences, Franklin Lakes, NJ), anti-TNC (Millipore), anti-Brevican (Abcam), and anti-laminin (Millipore).
To observe tumor cell migration in vivo, we mixed one portion of red fluorescent protein (RFP)-labeled control non-silencing cells (50,000) with three portion of green fluorescent protein (GFP)-labeled UGDH knockdown cells (150,000), and injected the mixture into the same mouse brains (n=3). Animals were sacrificed after 4 weeks and brain sections were observed under fluorescence microscopy.
To observe tumor cell migration in vivo, we mixed one portion of red fluorescent protein (RFP)-labeled control non-silencing cells (50,000) with three portion of green fluorescent protein (GFP)-labeled UGDH knockdown cells (150,000), and injected the mixture into the same mouse brains (n=3). Animals were sacrificed after 4 weeks and brain sections were observed under fluorescence microscopy.
4
Imaging Nanoparticle-Tumor Interactions
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Tumours collected for peptide and IO-NP binding and post-MRI were analysed. Tissue distribution of fluorescein (FAM)-labelled CSG, FAM-CREKA, FAM-IO-NP, FAM-CSG-IO-NP or FAM-CREKA-IO-NP were detected on 8 μm tissue cross-sections based on their fluorescence intensity and reactivity to anti-fluorescein-HRP antibody (polyclonal, GeneTex) and counter-stained with haematoxylin or methyl green (Vector Laboratories, Burlingame, CA, USA). For co-staining analysis, the following antibodies were used: anti-CD31 (390; ebioscience), anti-laminin (polyclonal; Millipore) and anti-collagen I (polyclonal; Abcam). For secondary detection, fluorescence-labelled, 594-conjugated anti-rat, rabbit or goat IgG (Life Technologies, Carlsbad, CA, USA) were used. Images were captured on a Nikon Ti-E microscope (Nikon Instrument Inc., Melville, NY, USA) or ScanScope XT (Aperio Technology, Inc., Vista, CA, USA). Image analysis and quantification were performed either using NIS software modules (version 4.0) or ImageScope version 12.1.0.5029 (Aperio Technology, Inc., Vista, CA, USA).
5
Immunostaining of Tibialis Anterior Muscle Sections
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Tibialis anterior (TA) muscle sections were prepared for staining essentially as previously described6 (link),48 (link). For OSMR staining, whole TA muscles were first fixed with 0.5% paraformaldehyde/PBS for 2 h at 4 C, followed by incubation in 20% sucrose/PBS overnight at 4 C. 10 μM sections were cut for all stainings. Antibodies were: anti-laminin (Millipore, catalog #05−206, 1:250), anti-GFP (Invitrogen, catalog #A11122, 1:200), anti-OSMR (R&D, catalog #AF665, 1:200), anti-Pax7 (Santa Cruz Biotechnology, catalog #sc81648, 1:50; or Developmental Studies Hybridoma Bank, 2 μg/mL final concentration), AlexaFluor 594–conjugated donkey anti-rat IgG1 and AlexaFluor 488–conjugated donkey anti-rabbit (Jackson ImmunoResearch, catalog # 712-585-150 and 711-545-152 respectively, 1:200 each). Nuclei were counterstained with either DAPI (Invitrogen) or TO-PRO-3 (Invitrogen). Images were acquired with an AxioPlan2 epifluorescent microscope (Carl Zeiss) with ORCA-ER digital camera (Hamamatsu Photonics).
6
Immunohistochemical Analysis of Donor-Cell Transgene
7
Glioblastoma Xenograft Model in SCID Mice
8
Quantifying Activated MuSCs in Transplanted Muscle
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
We collected the culture progeny of MuSCs from aged Myf5nLacZ/+/Luciferase double-transgenic mice23 (link),43 (link) by incubation with 0.1% trypsin in PBS for 2 min at 37 °C and transplanted them into tibialis anterior muscles of hindlimb-irradiated NOD/SCID mice. One month after transplant, we injected notexin to damage recipient muscles and activate MuSCs in vivo. Four days later, we collected, fixed, and cryosectioned recipient muscles, as described above. We performed immunohistological analysis of transverse tissue sections to detect β-galactosidase+ cells (indicating a donor-derived cell expressing Myf5, a marker of MuSC activation) in the satellite cell position within the myofiber basal lamina, as defined by laminin staining. We stained sections with anti-Laminin (Millipore, clone A5, catalog # 05-206, 1:250) and anti-β-galactosidase (Invitrogen, catalog # A11132, 1:100) primary antibodies and then with appropriate secondary antibodies (Invitrogen). We counter-stained nuclei with Hoechst 33342 (Invitrogen). We acquired images with an AxioPlan2 epi-fluorescent microscope (Carl Zeiss) with Plan NeoFluar 10×/0.30NA or 20×/0.75NA objectives (Carl Zeiss) and an ORCA-ER digital camera (Hamamatsu). We captured digital images in OpenLab software (Improvision) and assembled them using Photoshop software (Adobe) with consistent contrast adjustments across all images.
9
Immunohistochemical Analysis of Tissue Sections
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Samples fixed in Bouin solution and embedded in paraffin were serially sectioned at 6 μm. Sections were deparaffinated, rehydrated and heat-induced epitope retrieval was performed using sodium citrate buffer (10 mM sodium citrate, 0.05% Tween-20, pH 6) at 95 °C for 20 min. The procedure was conducted using the HRP/DAB Detection IHC Kit (ab64264, Abcam, Cambridge, UK). Incubation with the following primary antibodies was performed at 4 °C overnight: anti-E-cadherin (ab152102, Abcam), anti-laminin (L9393, Sigma), anti-PCNA (HPA030521, Sigma). Mayer’s hematoxylin was used as a counterstain. The images were taken with a Nikon Eclipse E600 light microscope (Amstelveen, The Netherlands).
10
Investigating the Role of LSD1 and FOXK1 in Muscle Metabolism
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Dex and insulin (from bovine pancreas) were purchased from Sigma-Aldrich, and STZ was from Wako. An LSD1 inhibitor, T-3775440 hydrochloride, was from MedChemExpress. The primary antibodies used for western blot, co-IP, immunohistochemistry, and immunocytochemistry experiments were as follows: anti-LSD1 (Abcam, ab17721), anti-FOXK1 (Abcam, ab18196), anti-ERRγ (Abcam, ab12893), anti-MHC type I (Developmental Studies Hybridoma Bank [DHSB], BA-F8), anti-MHC type IIA (DHSB, SC-71), anti-Laminin (Sigma-Aldrich, L9393), anti-Akt (Cell Signaling Technology [CST], #9272), anti-Phospho-Akt (Ser473) (CST, #9271), anti-4E-BP1 (CST, #9454), anti-Phospho-4E-BP1 (Thr37/46) (CST, #2855), anti-SQSTM1(p62) (Santa Cruz, sc-28359), anti-GAPDH (Santa Cruz, sc-25778), anti-LC3(MBL, PM036), and anti-Sin3A (Santa Cruz, sc-994). The secondary antibodies used were as follows: anti-mouse IgG-horseradish peroxidase (GE Healthcare, NA931V), anti-rabbit IgG-horseradish peroxidase (GE Healthcare, NA934V or CST, #7074), Alexa Fluor 488 anti-Mouse IgG1 (Thermo Fisher, A21121), Alexa Fluor 350 anti-Mouse IgG2b (Thermo Fisher, A21140), and Cy3 anti-Rabbit IgG (Jackson ImmunoResearch, 711-165-152). The antibodies used for ChIP experiments were anti-tri-methylated histone H3K4 (Millipore, 07-473), anti-pan histone H3 (Abcam, ab1791), and normal rabbit IgG (Santa Cruz, sc-2027).
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!