M. smegmatis was fixed in 2% paraformaldehyde/2.5% glutaraldehyde (Polysciences Inc.) in 100 mM sodium cocadylate buffer, pH 7.2 for 1 hour at room temperature. Samples were washed in sodium cacodylate buffer and postfixed in 1% osmium tetroxide (Polysciences Inc.) for 1 hr. Samples were then rinsed extensively in dH20 prior to en bloc staining with 1% aqueous uranyl acetate (Ted Pella Inc.) for 1 hr. Following several rinses in dH20, samples were dehydrated in a graded series of ethanol and embedded in Eponate 12 resin (Ted Pella Inc.). Sections of 95 nm were cut with a Leica Ultracut UCT ultramicrotome (Leica Microsystems Inc.), stained with uranyl acetate and lead citrate, and viewed on a JEOL 1200 EX transmission electron microscope (JEOL USA Inc.) equipped with an AMT 8 megapixel digital camera and AMT Image Capture Engine V602 software (Advanced Microscopy Techniques).
Paraformaldehyde 2.5 glutaraldehyde
Manufactured by Polysciences
Sourced in United States
Paraformaldehyde/2.5% glutaraldehyde is a fixative solution used in laboratory settings. It is a mixture of paraformaldehyde and glutaraldehyde, which are chemical compounds used to preserve and fix biological samples for microscopy and other analytical techniques.
Automatically generated - may contain errors
Lab products found in correlation
Eponate 12 resin, by Ted Pella (26 mentions)
Osmium tetroxide, by Polysciences (26 mentions)
Aqueous uranyl acetate, by Ted Pella (24 mentions)
1200 ex transmission electron microscope, by JEOL (24 mentions)
Ultracut uct ultramicrotome, by Leica (18 mentions)
Amt 8 megapixel digital camera, by Advanced Microscopy Techniques (17 mentions)
Amt image capture engine v602, by Advanced Microscopy Techniques (15 mentions)
Uranyl acetate, by JEOL (7 mentions)
Ultracut uc7 ultramicrotome, by Leica (3 mentions)
Amt eight mega pixel digital camera, by Advanced Microscopy Techniques (2 mentions)
Uranyl acetate, by Leica (2 mentions)
Centrifuge 5810r, by Eppendorf (1 mentions)
Axio observer d1, by Zeiss (1 mentions)
Ultracut uct 7 ultramicrotome, by Leica (1 mentions)
H 7500 transmission electron microscope, by Hitachi (1 mentions)
Tannic acid, by Electron Microscopy Sciences (1 mentions)
26 protocols using paraformaldehyde 2.5 glutaraldehyde
1
Transmission Electron Microscopy of Mycobacterium smegmatis
2
S. aureus Cell Ultrastructure Analysis
3
Ultrastructural Analysis Protocol
4
Ultrastructural Analysis of Cells
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For ultrastructural analyses, cells were fixed in 2% paraformaldehyde/2.5% glutaraldehyde (Polysciences, Inc., Warrington, PA, USA) in 100 mM sodium cacodylate buffer, pH 7.2, for 2 h at room temperature and then overnight at 4°C. Samples were washed in sodium cacodylate buffer and post-fixed in 1% osmium tetroxide (Polysciences, Inc.) for 1 h at room temperature. Samples were then rinsed extensively in dH2O prior to en bloc staining with 1% aqueous uranyl acetate (Ted Pella, Inc., Redding, CA, USA) for 1 h. Following several rinses in dH2O, samples were dehydrated in a graded series of ethanol and embedded in Eponate 12 resin (Ted Pella, Inc.). Sections of 95 nm were cut with a Leica Ultracut UC7 ultramicrotome (Leica Microsystems, Inc., Bannockburn, IL, USA), stained with uranyl acetate and lead citrate, and viewed on a JEOL 1200 EX transmission electron microscope (JEOL USA, Inc., Peabody, MA, USA) equipped with an AMT 8-megapixel digital camera and AMT Image Capture Engine V602 software (Advanced Microscopy Techniques, Woburn, MA, USA).
5
Ultrastructural Analysis of Biological Samples
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For ultrastructural analyses, samples were fixed in 2% paraformaldehyde-2.5% glutaraldehyde (Polysciences Inc., Warrington, PA) in 100 mM sodium cacodylate buffer, pH 7.2, for 2 h at room temperature and then overnight at 4°C. Samples were washed in sodium cacodylate buffer at room temperature and postinfection fixed in 1% osmium tetroxide (Polysciences Inc.) for 1 h. Samples were then rinsed extensively in distilled water (dH2O) prior to en bloc staining with 1% aqueous uranyl acetate (Ted Pella, Redding, CA) for 1 h. Following several rinses in dH2O, samples were dehydrated in a graded series of ethanol and embedded in Eponate 12 resin (Ted Pella Inc.). Sections of 95 nm were cut with a Leica Ultracut UCT ultramicrotome (Leica Microsystems Inc., Bannockburn, IL), stained with uranyl acetate and lead citrate, and viewed on a JEOL 1200 EX transmission electron microscope (JEOL USA Inc., Peabody, MA) equipped with an AMT 8-megapixel digital camera and AMT Image Capture Engine V602 software (Advanced Microscopy Techniques, Woburn, MA) as part of the Microbiology Imaging Facility, Washington University in St. Louis.
6
Transmission Electron Microscopic Analysis of Streptococcus mutans
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Transmission electron microscopic (TEM) analysis was carried out similarly as described previously [29 (link)]. Briefly, S. mutans strains were grown in BHI until mid-exponential phase, harvested by centrifugation at 4,000 rpm, 4°C for 10 minutes, washed twice with PBS, and then fixed in 2% paraformaldehyde/2.5% glutaraldehyde (Polysciences, Warrington, PA) in PBS for 1 h at room temperature. Cells were washed in PBS and post-fixed in 1% osmium tetroxide (Polysciences) for 1 h. Samples were then rinsed extensively in dH20 prior to en bloc staining with 1% aqueous uranyl acetate (Ted Pella Inc., Redding, CA) for 1 hour. The cells were then washed with dH20 and dehydrated in a graded series of ethanol and embedded in Eponate 12 resin (Ted Pella Inc., Redding CA). Sections of 90–100 nm were prepared, stained with uranyl acetate and lead citrate, and viewed under a JEOL 1200 EX transmission electron microscope (JEOL USA Inc., Peabody, MA).
7
Ultrastructural Analysis of Biological Samples
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For ultrastructural analyses, samples were fixed in 2% paraformaldehyde/2.5% glutaraldehyde (Polysciences, Warrington, PA) in 100 mM sodium cacodylate buffer, pH 7.2 for 2 hours at room temperature and then overnight at 4°C. Samples were washed in sodium cacodylate buffer at room temperature and postfixed in 1% osmium tetroxide (Polysciences) for 1 hour. Samples were then rinsed extensively in dH20 prior to en bloc staining with 1% aqueous uranyl acetate (Ted Pella, Redding, CA) for 1 hour. Following several rinses in dH20, samples were dehydrated in a graded series of ethanol and embedded in Eponate 12 resin (Ted Pella). Sections of 95 nm were cut with a Leica Ultracut UCT ultramicrotome (Leica Microsystems, Bannockburn, IL), stained with uranyl acetate and lead citrate, and viewed on a JEOL 1200 EX transmission electron microscope (JEOL USA, Peabody, MA) equipped with an AMT 8-MP digital camera and AMT Image Capture Engine V602 software (Advanced Microscopy Techniques, Woburn, MA).
8
Biofilm Ultrastructural Analysis by TEM
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Biofilm grown on urinary catheter surface was processed for structure characterization using transmission electron microscopy (TEM) (84 (link)). For microstructural characterization, 1 cm catheter with biofilm formed on the inside surface was fixed in 2% paraformaldehyde/2.5% glutaraldehyde (Polysciences Inc.) in 100 mM sodium cacodylate buffer (pH 7.2) for 1 hour at room temperature. Samples were washed in sodium cacodylate buffer and postfixed in 1% osmium tetroxide (Polysciences Inc.) for 1 hour. Samples were then rinsed extensively in deionized water prior to en bloc staining with 1% aqueous uranyl acetate (Ted Pella Inc.) for 1 hour. Following several rinses in deionized water, samples were dehydrated in a graded series of ethanol and embedded in Eponate 12 resin (Ted Pella Inc.). Sections of 95 nm were cut with a Leica Ultracut UCT ultramicrotome (Leica Microsystems Inc.), stained with uranyl acetate and lead citrate, and viewed on a JEOL 1200 EX transmission electron microscope (JEOL USA Inc.) equipped with an AMT 8 megapixel digital camera and AMT Image Capture Engine V602 software (Advanced Microscopy Techniques).
9
EMCV Infection in A549 Cells
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EMCV was incubated with A549 cells (wild type, TNK2 KO and WASL KO) at an MOI of 20 for 1 hr on ice. Cells were then washed with serum free DMEM and incubated with A549 culture media (2% FBS) for 6 hr. Infected cells were washed with PBS and fixed with 2% paraformaldehyde, 2.5% glutaraldehyde (Polysciences Inc, Warrington, PA) in 100 mM cacodylate buffer for 1 hr at room temperature. Next, cells were scraped from plates using a rubber cell scraper and cell pellets were embedded in agarose. Agarose embedded cell pellets were post-fixed in 1% osmium tetroxide (Polysciences Inc) for 1 hr, then rinsed extensively in dH20 prior to en bloc staining with 1% aqueous uranyl acetate (Ted Pella Inc, Redding, CA) for 1 hr. Following several rinses in dH20, samples were dehydrated in a graded series of ethanol and embedded in Eponate 12 resin (Ted Pella Inc). Sections of 95 nm were cut with a Leica Ultracut UCT ultramicrotome (Leica Microsystems Inc, Bannockburn, IL), stained with uranyl acetate and lead citrate, and viewed on a JEOL 1200EX transmission electron microscope (JEOL USA, Peabody, MA) equipped with an AMT eight mega-pixel digital camera (Advanced Microscopy Techniques, Woburn, MA).
10
Detailed Imaging Protocols for Parasite Detection
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Parasites monitored by
thin smear were dyed using Harleco Hemacolor stains (MilliporeSigma).
Images were taken using a Zeiss Axio Observer.D1 at the Washington
University Molecular Microbiology Imaging Facility. For transmission
electron microscopy, infected RBCs were fixed in 2% paraformaldehyde/2.5%
glutaraldehyde (Polysciences Inc., Warrington, PA) in 100 mM sodium
cacodylate buffer, pH 7.2 for 1 h at room temperature. Samples were
washed in sodium cacodylate buffer at room temperature and postfixed
in 1% osmium tetroxide (Polysciences Inc.) for 1 h. Samples were rinsed
extensively in deionized water prior to en bloc staining
with 1% aqueous uranyl acetate (Ted Pella Inc., Redding, CA) for 1
h. Following several rinses in deionized water, samples were dehydrated
in a graded series of ethanol and embedded in Eponate 12 resin (Ted
Pella Inc.). Sections of 95 nm were cut with a Leica Ultracut UCT
ultramicrotome (Leica Microsystems Inc., Bannockburn, IL), stained
with uranyl acetate and lead citrate, and viewed on a JEOL 1200 EX
transmission electron microscope (JEOL USA Inc., Peabody, MA) equipped
with an AMT 8-megapixel digital camera and AMT Image Capture Engine
V602 software (Advanced Microscopy Techniques, Woburn, MA).
thin smear were dyed using Harleco Hemacolor stains (MilliporeSigma).
Images were taken using a Zeiss Axio Observer.D1 at the Washington
University Molecular Microbiology Imaging Facility. For transmission
electron microscopy, infected RBCs were fixed in 2% paraformaldehyde/2.5%
glutaraldehyde (Polysciences Inc., Warrington, PA) in 100 mM sodium
cacodylate buffer, pH 7.2 for 1 h at room temperature. Samples were
washed in sodium cacodylate buffer at room temperature and postfixed
in 1% osmium tetroxide (Polysciences Inc.) for 1 h. Samples were rinsed
extensively in deionized water prior to en bloc staining
with 1% aqueous uranyl acetate (Ted Pella Inc., Redding, CA) for 1
h. Following several rinses in deionized water, samples were dehydrated
in a graded series of ethanol and embedded in Eponate 12 resin (Ted
Pella Inc.). Sections of 95 nm were cut with a Leica Ultracut UCT
ultramicrotome (Leica Microsystems Inc., Bannockburn, IL), stained
with uranyl acetate and lead citrate, and viewed on a JEOL 1200 EX
transmission electron microscope (JEOL USA Inc., Peabody, MA) equipped
with an AMT 8-megapixel digital camera and AMT Image Capture Engine
V602 software (Advanced Microscopy Techniques, Woburn, MA).
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