For immunofluorescence analysis, 25 μm serial cryosections were obtained using a CM3050 cryostat (Leica Microsystems, Wetzlar, Germany). After incubation in a blocking buffer (1 × PBS/1% bovine serum albumin/0.3% Triton X-100) for 1 hr at room temperature, sections were incubated with the following primary antibodies overnight in PBS at 4°C: mBDNF (ab75040, Abcam), pTrkB (ab131483, Abcam), TH (sc25269, Santa Cruz Biotechnology, Inc), DAT (sc32259, Santa Cruz Biotechnology, Inc), BrdU (MCA2483, Bio-Rad, Hercules, CA, USA), NeuN (ABN78, Millipore Corporation), and Iba1 (019–19741, Wako Chemicals, Richmond, VA, USA). Sections were washed with PBST and incubated with fluorescent secondary antibodies (A11001, A11007, or A11037, Invitrogen, Carlsbad, CA, USA; CA94010, Vector Laboratories, Inc, Burlingame, CA, USA) for 2 hr at room temperature. Sections were mounted onto slides using a mounting medium (H-1200, Vector Laboratories, Inc) and imaged using a fluorescence microscope (Carl Zeiss Imager M1, Carl Zeiss AG, Oberkochen, Germany). Immunofluorescence was analyzed using the IMT i-Solution Inc 10.1 (17TH-5989 Walter Gage Rd., Vancouver, BC, CA) image analysis software.
Carl imager m1
Manufactured by Zeiss
Sourced in Germany
The Carl Zeiss Imager M1 is a high-precision optical microscope designed for advanced imaging and analysis applications. It features a robust and ergonomic design, providing reliable performance and versatile functionality. The Imager M1 is equipped with state-of-the-art optics and technologies to ensure accurate and detailed imaging results.
Automatically generated - may contain errors
Lab products found in correlation
H 1200, by Vector Laboratories (2 mentions)
Mounting medium, by Vector Laboratories (2 mentions)
Ab75040, by Abcam (1 mentions)
Cm3050 cryostat, by Leica (1 mentions)
A11037, by Thermo Fisher Scientific (1 mentions)
A11007, by Thermo Fisher Scientific (1 mentions)
Neuronal nuclei (neun), by Merck Group (1 mentions)
Sc 25269, by Santa Cruz Biotechnology (1 mentions)
Ca 94010, by Vector Laboratories (1 mentions)
A11001, by Thermo Fisher Scientific (1 mentions)
Primary anti mmp 9, by Merck Group (1 mentions)
Goat anti rabbit igg tr, by Vector Laboratories (1 mentions)
Anti mmp 3, by Santa Cruz Biotechnology (1 mentions)
Ab40390, by Abcam (1 mentions)
Mounting medium, by Agilent Technologies (1 mentions)
Mca2483, by Bio-Rad (1 mentions)
Carboxy h2dcfda, by Thermo Fisher Scientific (1 mentions)
Ab79351, by Abcam (1 mentions)
6 protocols using carl imager m1
1
Immunofluorescence Analysis of Neural Markers
2
In Situ Zymography for Gelatinase and Collagenase
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
To detect the activity and location of gelatinase and collagenase, in situ zymography was performed. Dorsal skin tissues were rapidly frozen using 2-methylbutane and liquid nitrogen, and 20-μm cryosections were prepared. Sections were reacted at 37 °C for approximately 8 h in dark humidified chambers with a Molecular Probes EnzChek Gelatinase/Collagenase Assay Kit (Life Technologies, Eugene, OR, USA). After washing, the skins were fixed in 10% neutral-buffered formalin in the dark. To evaluate MMP-9 and MMP-3 co-localization, sections were incubated with primary anti-MMP-9 (Millipore, Billerica, MA, USA) or anti-MMP-3 (Santa Cruz Biotechnology Inc., CA, USA) in antibody dilution buffer (1 × PBS, 1% bovine serum albumin [BSA], and 0.3% Triton X-100) overnight in the dark, followed by washing with PBS. The sections were incubated with the corresponding secondary goat anti-rabbit IgG-TR (Vector Laboratories Inc.) or rabbit anti-goat IgG-TR (Vector Laboratories Inc.) antibody for 2 h at room temperature in the dark, followed by washing with PBS. Subsequently, the slides were mounted using mounting medium (Vector Laboratories Inc.), and images were acquired using a fluorescence microscope (Carl Zeiss Imager M1, Carl Zeiss Inc.). IMT i-Solution (IMT i-Solution Inc.) was used for automatic measurement of the integrated optical density (IOD).
3
Brain Tissue Fixation and Immunostaining
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
After behavioral assessments, the rats anesthetized with isoflurane received intracardial perfusion with saline, followed by 4% paraformaldehyde in PBS. The brains were removed, post-fixed in the same fixative for 4 h at 4°C, and immersed in 30% sucrose for 48 h for cryoprotection. Frozen 20 μm-thick sections from bregma 0 to +1 mm were incubated for post-fixation with formaldehyde, and blocked with a blocking buffer (1X PBS/5% normal goat serum/0.3% Triton X-100) for 1 h at room temperature. The sections were incubated with the following primary antibodies to BrdU (MCA2483, AbD Serotec, Hercules, CA, USA), NeuN (ABN78, EMD; Millipore) and MBP (ab40390; Abcam) overnight in PBS at 4°C. After washing with PBS, the sections were incubated with the fluorescent secondary antibodies against horse anti-mouse (FI-2000), goat anti-rabbit (TI-1000) and goat anti-rabbit (FI-1000) (all from Vector Laboratories, Inc., Burlingame, CA, USA) for 2 h at room temperature in the dark, and then washed 3 times with PBS. Subsequently, slides were mounted in mounting medium (Dako, Glostrup, Denmark) and captured using a fluorescence microscope (Carl Zeiss Imager M1; Carl Zeiss, Inc., Gottingen, Germany). IMT i-Solution (IMT i-Solution Inc., Burnaby, BC, Canada) was used for the automatic measurement of cell counting, thickness and integrated optical density (IOD).
4
Glutamate-Induced Oxidative Stress Assay
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
HT22 cells were cultured in 96-well white plates at a density of 5x103 cells per well. After adherence, cells were pretreated with PMC-12 for 24 h and then exposed to 5 mM glutamate for 24 h. Treated cells were washed with PBS. Carboxy-H2DCFDA (20 μM) (Invitrogen) was applied to the cells, followed by incubation for 1 h in a 37 °C incubator. Fluorescence was measured using a Mutilabel counter (Perkin Elmer 1420, MA, USA). Accumulation of intracellular ROS was observed and photographed under a fluorescence microscope (Carl Zeiss Imager M1, Carl Zeiss, Inc., Gottingen, Germany). In addition, cells were harvested, resuspended in 1 ml PBS with 20 μM carboxy-H2DCFDA (Invitrogen), and then incubated for 1 h at 37 °C. After washing, cellular fluorescence was measured using a flow cytometer.
5
Immunofluorescence Profiling of Murine Brain
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Coronal serial cryosections of both hemispheres, each 25-μm-thick, were obtained. The cryosections were post-fixed with a 4% paraformaldehyde solution for 15 minutes before being washed with phosphate-buffered saline (PBS). Following that, the samples were processed using a standard immunofluorescence methodology for cell signaling studies, which involved blocking the samples with antibody dilution buffer (1× PBS with 5% normal goat serum and 0.3% Triton X-100). The following primary antibodies were used to stain the samples at 4℃ overnight: BrdU (OBT0030G, AbD Serotec, Oxford, UK), Dcx (Ab18723, Abcam, Cambridge, UK), NeuN (MAB377 and ABN78, Millipore Corporation, Billerica, MA, USA), GFAP (Z0334, DAKO, Santa Clara, CA, USA), and Sox2 (ab79351, Abcam). The following day, after three washes with PBS with Tween 20 (PBST), the samples were incubated with secondary antibodies for 2 hours. The stained brain tissues were mounted using a solution of 4’,6-diamidino-2-phenylindole (DAPI; H-1200, Vector Laboratories, Inc., Burlingame, CA, USA). Five brain slices and 10× magnified images of coronal brain sections were selected for cell counting. The number of cells was calculated using i-solution (IMT i-solution Inc.) after capturing images with a fluorescence microscope (Carl Zeiss Imager M1, Carl Zeiss AG, Oberkochen, Germany).
6
Fluoromyelin-Based Myelin Quantification
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Mouse brain was removed and post-fixed in 4% paraformaldehyde in PBS for 24 h at 4 °C before cryoprotection in 30% sucrose. Frozen 20-μm sections were rinsed in PBS for at least 20 min. Sections were then incubated with FluoroMyelinTM Green fluorescent myelin stain (1:300, Molecular probes, Eugene, OR, USA) for 20 min at room temperature, and then slides were mounted using mounting medium (Vector Laboratories, Inc., Burlingame, CA, USA). Semi-quantification of the intensity of fluoromyelin staining was conducted using 10× magnification images that were captured using a fluorescence microscope (Carl Zeiss Imager M1, Carl Zeiss, Inc., Gottingen, Germany).
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!