Huh7 YFP-TIA1 cells (105) were seeded in a 35-mm dish with 20-mm cover glass no. 1.5 (MatTek). Confocal Z-stacks (200 nm Z spacing) of whole cells were acquired using the YFP-TIA1 signal (514 nm excitation) on a Nikon Ti Eclipse using an Apo total internal reflection fluorescence (TIRF) 60× oil (NA 1.49) objective (Nikon) with a digital image using the Orca Flash 4 v2 camera (6.5 × 6.5 μm pixel size, Hamamatsu). Substacks with 1-μm Z spacing including the whole cell volume were chosen and manually circled to measure area per stack. Total cell volume was calculated by multiplication of total area volume with 1-μm stack thickness.
Orca flash 4 v2 camera
Manufactured by Hamamatsu Photonics
The Orca Flash 4 v2 is a scientific-grade digital camera produced by Hamamatsu Photonics. It features a sCMOS image sensor with a resolution of 2048 x 2048 pixels and a pixel size of 6.5 μm. The camera is capable of capturing images at a maximum frame rate of 100 frames per second.
Automatically generated - may contain errors
2 protocols using orca flash 4 v2 camera
1
Quantifying 3D Cell Volume Imaging
2
Three-Dimensional Localization Microscopy of Bacteria
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Three-dimensional d-STORM
imaging was performed using Vutara SR352 microscope (Bruker, USA)
based on single-molecule localization biplane technology. Images were
recorded using 1.3 NA 60× silicon oil immersion objective (Olympus)
and Hamamatsu Orca Flash 4v2 camera with a frame rate at 50 Hz. Bacteria
were incubated with liposomes labeled with DiR (0.5% mol/mol) dye
for 4 h, then washed three with PBS. Next, bacteria were stained with
1 mg/mL membrane stain FM1-43 (Thermo Fisher Scientific T35356), washed
three times with PBS, centrifuged at 5000 rpm for 2 min at 25 °C,
resuspended. The bacteria were fixed with 2.8% formaldehyde (FA),
0.04% glutaraldehyde-formaldehyde (GA) for 15 min, washed three times,
centrifuged at 5000 rpm for 2 min at 25 °C, and resuspended in
10 μL PBS. Bacteria were dropped on a poly-l -lysine
(Sigma P8920) coated MatTek plate (P35G-1.5-7-C), and imaging was
performed in the presence of an imaging buffer (7 μM glucose
oxidase (Sigma), 56 nM catalase (Sigma), 2 mM cysteamine (Sigma),
50 mM Tris, 10 mM NaCl, 10% glucose, pH 8). Subsequently, STORM images
of liposomes were recorded using 640 nm excitation laser (maximal
excitation of 6 kW/cm2) with a collection of 3000 frames.
Data were analyzed with the Vutara SRX 7.0.00rc24 software.
imaging was performed using Vutara SR352 microscope (Bruker, USA)
based on single-molecule localization biplane technology. Images were
recorded using 1.3 NA 60× silicon oil immersion objective (Olympus)
and Hamamatsu Orca Flash 4v2 camera with a frame rate at 50 Hz. Bacteria
were incubated with liposomes labeled with DiR (0.5% mol/mol) dye
for 4 h, then washed three with PBS. Next, bacteria were stained with
1 mg/mL membrane stain FM1-43 (Thermo Fisher Scientific T35356), washed
three times with PBS, centrifuged at 5000 rpm for 2 min at 25 °C,
resuspended. The bacteria were fixed with 2.8% formaldehyde (FA),
0.04% glutaraldehyde-formaldehyde (GA) for 15 min, washed three times,
centrifuged at 5000 rpm for 2 min at 25 °C, and resuspended in
10 μL PBS. Bacteria were dropped on a poly-
(Sigma P8920) coated MatTek plate (P35G-1.5-7-C), and imaging was
performed in the presence of an imaging buffer (7 μM glucose
oxidase (Sigma), 56 nM catalase (Sigma), 2 mM cysteamine (Sigma),
50 mM Tris, 10 mM NaCl, 10% glucose, pH 8). Subsequently, STORM images
of liposomes were recorded using 640 nm excitation laser (maximal
excitation of 6 kW/cm2) with a collection of 3000 frames.
Data were analyzed with the Vutara SRX 7.0.00rc24 software.
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!