Microscopic observation was conducted to study the influence of LED lights on micromorphology of hairy roots. Samples were prepared using the method reported previously (Gai et al. 2020 (link)). A DM 4000B fluorescence microscope (Leica, Germany) equipped with a DS-Ri1 digital camera (Nikon, Japan) was used to acquire images under the magnification of ×10.
Dm4000b fluorescence microscope
Manufactured by Leica camera
Sourced in Germany
The DM4000B is a fluorescence microscope designed for advanced microscopy applications. It features a high-performance optical system and a variety of illumination options to enable detailed observation and analysis of samples. The DM4000B is suitable for a range of research and laboratory tasks requiring advanced microscopy techniques.
Automatically generated - may contain errors
Lab products found in correlation
Lsm 510 meta confocal microscope, by Zeiss (2 mentions)
Ds ri1 digital camera, by Nikon (1 mentions)
Alexa fluor 594, by Abcam (1 mentions)
Anti caspase 3 antibody, by Abcam (1 mentions)
Trypan blue solution, by Merck Group (1 mentions)
Vector red alkaline phosphatase substrate kit 1, by Vector Laboratories (1 mentions)
Prolong gold antifade reagent, by Thermo Fisher Scientific (1 mentions)
Ab167453, by Abcam (1 mentions)
Chamber slide, by Thermo Fisher Scientific (1 mentions)
Protein block, by Agilent Technologies (1 mentions)
Prolong gold antifade mounting medium, by Thermo Fisher Scientific (1 mentions)
Ab137321, by Abcam (1 mentions)
Ab109185, by Abcam (1 mentions)
D3571, by Thermo Fisher Scientific (1 mentions)
Ab735, by Proteintech (1 mentions)
Sc 71064, by Santa Cruz Biotechnology (1 mentions)
Sc 398898, by Abcam (1 mentions)
Glycerinum, by Merck Group (1 mentions)
Freezing microtome, by Leica camera (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (1 mentions)
16 protocols using dm4000b fluorescence microscope
1
LED Effects on Hairy Root Micromorphology
2
Apoptosis Induction in PSMA-Targeted Tumor Therapy
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Animals bearing PC3pip tumors were divided into 7 groups: (1) mice receiving PBS; (2) mice receiving 100 nmol/kg PSMA-1-MMAE-IR700 with PDT; (3) mice receiving equal doses of PSMA-1-MMAE-IR700 to group 2, but not receiving PDT; (4) Mice receiving equal doses of PSMA-1-IR700 to group 2 with PDT treatment; (5) Mice receiving equal doses of PSMA-1-IR700 to group 2 without PDT; (6) mice receiving equal dose of PSMA-1-IR700 to group 2 + free MMAE with PDT (MMAE normalized to that delivered by PSMA-1-MMAE-IR700); and (7) mice receiving equal dose of PSMA-1-IR700 to group 2 + free MMAE without PDT. Animals were treated with one single dose and were sacrificed at 4-day post treatment. Tumors were snap-frozen in OCT, cut into 10 μm thick sections and fixed on slides. Induction of apoptosis by the treatment was determined by rabbit polyclonal anti-Caspase-3 antibody (Abcam, Cambridge, UK). A goat anti-rabbit polyclonal antibody labeled by Alexa Fluor-594 was used as secondary antibody (Abcam, Cambridge, UK). The presence of apoptosis was determined by fluorescence images under Leica DM4000B fluorescence microscope at 10 ×. H&E staining of tumor tissues was performed in adjacent sections to check the histology of the tumors. Experiments were repeated in 5 mice.
3
GFP Fixation and Microscopy Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cells were fixed according to the protocol described on the Koshland lab web site (http://mcb.berkeley.edu/labs/koshland/Protocols/MICROSCOPY/gfpfix.html ): collected cell pellets were resuspended in paraformaldehyde solution (4% paraformaldehyde, 3.4% sucrose) and incubated at room temperature for 15 min. Cells were then washed once with KPO4/sorbitol solution and resuspended in 50 μL KPO4/sorbitol solution (0.1 M KPO4 pH 7.5, 1.2 M sorbitol) and stored at 4°C until visualization. Before fluorescence microscopy, cells were sonicated and incubated with 0.5 μg/mL DAPI for at least an hour. Cells were visualized using a Leica DM4000 B fluorescence microscope and greater than 200 cells were scored for each sample. Multiple replicates performed for the DAPI anaphase entry experiments are presented in Figures 5 , 6 , and S6 .
4
Evaluating Cell Death in Root Tips
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
To assess cell death in the root tips under Cd stress, the roots were immersed in 4 mg ml-1 Trypan blue solution (Sigma–Aldrich, Saint Louis, MO, USA) for 15 min at room temperature and then washed with distilled water three times. The samples were then observed by light microscopy, and pictures were taken as detailed earlier.
For PI staining, the seedlings were immersed in the PI (Sigma–Aldrich, St. Louis, MO, USA) solution (final concentration of 1 μg ml-1) for 10 min at room temperature in the dark and then washed with phosphate buffer solution (PBS) (pH 7.4) three times. The samples were then examined using a DM 4000B fluorescence microscope (Leica, Germany) with an excitation wavelength of 546 nm. Both experiments were repeated three times. For each treatment and genotype, at least 20 roots were analyzed for both stains, and one representative image was selected for the figure.
For PI staining, the seedlings were immersed in the PI (Sigma–Aldrich, St. Louis, MO, USA) solution (final concentration of 1 μg ml-1) for 10 min at room temperature in the dark and then washed with phosphate buffer solution (PBS) (pH 7.4) three times. The samples were then examined using a DM 4000B fluorescence microscope (Leica, Germany) with an excitation wavelength of 546 nm. Both experiments were repeated three times. For each treatment and genotype, at least 20 roots were analyzed for both stains, and one representative image was selected for the figure.
5
GFP Fixation and Microscopy Protocol
6
Visualizing Calcified Aortic Adventitial Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Alkaline phosphatase–positive cells in aortic adventitial cell cultures were visualized as previously described, but using the Vector Red fluorescent substrate (14 (link),18 (link)). Briefly, aortic mesenchymal cells were cultured on a type I collagen–precoated four-chamber slide (60,000 cells/chamber) for 8 days. During the final 6 days, cells were treated with β-glycerophosphate 5 mmol/L and ascorbic acid 50 μg/mL. Cells were then washed three times with Tris-buffered saline (TBS) (20 mmol/L Tris HCl, 1.5 mol/L NaCl, pH 7.5), fixed with 4% paraformaldehyde in TBS for 4 min, and rinsed three times with TBS. Staining with Vector Red reagent (Vector Red Alkaline Phosphatase Substrate Kit I, catalog number SK-5100; Vector Laboratories) was performed for 1 h in the dark per the manufacturer’s protocol. After washing three times with TBS and twice with distilled water, nuclei were stained with DAPI (Prolong Gold Antifade Reagent, catalog number P36931; Molecular Probes) and the slides coverslipped. Photomicrographs of alkaline phosphatase–positive fluorescent cells (TX2 filter cube), nuclei (DAPI filter cube), and phase contrast images were captured by a Leica DM4000 B fluorescence microscope at ×200 magnification.
7
Visualization of RFP Expression in Edited Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cells (unmodified, transfected with pAAVS1-(CMV)-RFP-Puro-DNR, or edited with CRISPR/Cas9 to insert the CMV-RFP expression cassette into the AAVS1 locus) were grown on chamber slides (Thermo Fisher Scientific, Waltham, MA) to reach 60-70% confluency, and were then fixed with 4% paraformaldehyde. Immunofluorescence staining for RFP was performed by permeabilization with 0.2% Triton-X for 15 min, blocking with protein block (Dako, Santa Clara, CA) for 30 min at RT, and incubation at 4°C overnight with a rabbit polyclonal anti-RFP primary antibody (ab167453, Abcam, Cambridge, UK) at a dilution of 1:500. The slides were washed the following day and incubated with a goat anti-Rabbit AlexaFluor® 594-conjugated secondary antibody (A-11072, Thermo Fisher Scientific) at a dilution of 1:500. Finally, the nuclei were counterstained with DAPI (1:1000) and coverslips were mounted with ProLong Gold Antifade Mounting Medium (Thermo Fisher Scientific), followed by sealing with clear nail polish. The slides were imaged using a Leica DM4000B fluorescence microscope. A control slide, stained with the secondary antibody only, was included to assess the degree of non-specific binding and determine the appropriate exposure settings for image acquisition.
8
Quantitative Analysis of Lesion Area and Cell Counts
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Lesion area and total cell numbers were quantified using images acquired with a Leica DM4000B fluorescence microscope at 10× magnification. PLP stained lesions were outlined and the selected area quantified using the ImageJ/Fiji processing software. Cells of interest within the area were counted manually. The total number of cells normalized to 1 mm2 lesion area was calculated using the formula [(number of cells in lesion/total area of lesion)] × 105. Statistical analysis was performed using Microsoft excel and GraphPad Prism 5 Software. Individual groups were compared using the unpaired, two‐tailed Student's t‐test at a 95% confidence interval. A value of P <0.05 was considered statistically significant.
9
Immunofluorescence Analysis of Epithelial Markers
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
MEECs were grown to an appropriate density and fixed in 4% (w/v) paraformaldehyde for 15 min, permeabilized with 0.5% Triton-X-100 for 15 min, and blocked with 5% bovine serum albumin for one hour at room temperature. After blocking, cells were labeled with the primary antibodies (1:100, rabbit anti-EpCAM, Proteintech, 21050-1-AP; 1: 100, rabbit anti-Mucin1, Abcam, ab109185; 1:100, mouse anti-P63, Abcam, ab735; 1:100, rabbit anti-CD44, Proteintech, 15675-1-AP; 1:100, mouse anti-ERα, Santa Cruz Biotechnologies, sc-71064; 1:100, mouse anti-PR, Santa Cruz Biotechnologies, sc-398898; 1:100, rabbit anti-Vimentin, Abcam, ab137321), and the secondary antibodies (1:100, fluorescently labeled goat anti-mouse lgG-cy3, BA1031, Boster company, Wuhan, China) according to the manufacture’s protocol. DAPI (0.5 mg/ml, D3571, Thermo Fisher) was used to stain the nucleus. Then, the fluorescence was detected by Leica DM4000B fluorescence microscope.
10
Immunofluorescence Analysis of Corneal Tissues
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Human corneoscleral tissues were fixed in 4% paraformaldehyde (Beyotime Biotechnology) for 20 min, dehydrated in graded sucrose, embedded in an optimum-cutting-temperature compound (SAKURA), and preserved at -80°C. The embedded tissue was cut into 10-μm-thick sections with a freezing microtome (Leica). For immunofluorescence staining, cultured cells were seeded on coverslips and fixed with 4% paraformaldehyde for 20 min. Sections and paraformaldehyde-fixed LECs were permeabilized with PBS (Songon Biotech) containing 0.3% Triton X-100 (Sigma-Aldrich) for 10 min, blocked with PBS containing 3% BSA (BSA, Roche) for 1 h at room temperature, and incubated with primary antibodies diluted in 1% BSA and 0.1% Triton X-100 overnight at 4°C. Thereafter, the sections and cells were washed thrice in PBS, incubated with secondary antibodies for 1 h at room temperature, and then washed thrice in PBS. Nuclei were labeled with 4′,6-diamidino-2-phenylindole (DAPI; Invitrogen) for 5 min at room temperature, and then samples were washed thrice in PBS and mounted in 50% glycerinum (Sigma-Aldrich). Images were acquired using a Leica TCS SP8 confocal microscope and a Leica DM4000B fluorescence microscope. Fluorescent signals in three randomly selected areas were quantified using ImageJ software (National Institutes of Health, USA). Antibody information is provided in Table S2 .
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!