The use of Lab-Tek tissue culture Chamber/ Slides and Flaskette for processing cultured monolayers for electron microscopy

1983 ◽  
Vol 41 ◽  
pp. 626-627
Author(s):  
K. Chien ◽  
R.L. Van De Velde ◽  
R.C. Heusser ◽  
J.W. Said
Keyword(s):  
Tissue Culture ◽  
Glass Slide ◽  
Culture Chamber ◽  
Major Difficulty ◽  
Cell Monolayers ◽  
Plastic Resin ◽  
Fast Procedure ◽  
In Situ Embedding ◽  
Chamber Slide

In situ embedding is a preferable means for the ultrastructural study of cultured cell monolayers. However, the major difficulty is the separation of the polymerized plastic resin from the supporting substrates. This abstract demonstrates a simple, fast procedure for using existing products for cell culture and in situ embedding of monolayers. Epoxy block surfaces up to 10 cm2 can be easily separated from an untreated glass slide and sectioned vertically or horizontally for EM studies.The Lab-Tek tissue culture Chamber/Slides or Flaskette are constructed so that 1 to 8 chamber compartments or a single flask is attached to a regular glass slide by gaskets. Different cell lines or procedures can be performed on the same chamber/slide. When the plastic chambers and gasket are removed, the glass slide can be coverslipped and filed for reference. We use these products routinely for cell cultures. When an EM study is needed, we fix the cell monolayers in situ within the chambers or flask, dehydrate with ethanol, infiltrate and embed in an epoxy resin.

In Situ Embedding of Cell Monolayers on Untreated Glass Surfaces for Vertical and Horizontal Ultramicrotomy.

1980 ◽  
Vol 38 ◽  
pp. 644-645
Author(s):  
Kai Chien
Keyword(s):  
Organic Solvent ◽  
Light Microscopy ◽  
Cell Monolayer ◽  
Glass Slide ◽  
Plastic Substrates ◽  
Cell Monolayers ◽  
Glass Surfaces ◽  
In Situ Embedding ◽  
The Ideal

Untreated glass is the ideal supporting substrate for cell cul¬ture growth since it is extremely flat and smooth, transparent and insoluable in organic solvent. However, difficulties have been encountered in removing polymerized epoxy resin from a glass surface following in situ cell monolayer embedment. Vari¬ous techniques have been made to grow cell cultures on either coated glass surfaces or plastic substrates. The purpose of this abstract is to describe a heat separation technique which when used together with a newly designed embedding mold allows cell monolayers to be transferred from untreated coverglass or glass slide to pre-shaped tissue blocks. The resulting tissue block can be easily separated and used directly for orientation light microscopy prior to ultramicrotomy.

In Situ Embedding of Cell Monolayers Cultured on Plastic Surfaces for Electron Microscopy

1991 ◽  
Vol 66 (5) ◽  
pp. 269-272 ◽  
Author(s):  
A. Beatrice Murray ◽  
Helga Schulze ◽  
Elisabeth Blauw
Keyword(s):  
Electron Microscopy ◽  
Cell Monolayers ◽  
Plastic Surfaces ◽  
In Situ Embedding

Capillary hydraulic conductivity is decreased by nitric oxide synthase inhibition

1995 ◽  
Vol 268 (5) ◽  
pp. H1856-H1861 ◽  
Author(s):  
R. E. Rumbaut ◽  
M. K. McKay ◽  
V. H. Huxley
Keyword(s):  
Nitric Oxide ◽  
Hydraulic Conductivity ◽  
Guanylate Cyclase ◽  
Soluble Guanylate Cyclase ◽  
No Synthase ◽  
Capillary Water ◽  
Cell Monolayers ◽  
Study Support ◽  
Oxide Synthase

Nitric oxide (NO) has been reported to modulate microvascular permeability to solutes in whole organs, venules, and cultured endothelial cell monolayers. NO derived from L-arginine via NO synthase activates soluble guanylate cyclase in vascular smooth muscle and endothelial cells. While the effects of NO on capillary water permeability have not been characterized, other activators of guanylate cyclase, such as sodium nitroprusside and atrial natriuretic peptide, increase capillary hydraulic conductivity (Lp). We hypothesized that inhibition of NO synthase with the arginine analogue, NG-monomethyl-L-arginine (L-NMMA), would decrease Lp from control levels. Lp was assessed in situ in single perfused frog mesenteric capillaries, first during control conditions (Lcontrolp) and then during superfusion (Ltestp) with either L-NMMA, NG-monomethyl-D-arginine (D-NMMA), a biologically inert enantiomer, or L-NMMA and L-arginine. Superfusion with 1 microM L-NMMA caused a decrease in Lp (Ltestp/Lcontrolp = 0.6 +/- 0.1, P < 0.001), whereas 1 microM D-NMMA was without effect on Lp (Ltestp/Lcontrolp = 1.0 +/- 0.2). The decrease in Lp by 1 microM L-NMMA was not only prevented by the presence of excess L-arginine (100 microM), but Lp increased from control (Ltestp/Lcontrolp = 1.4 +/- 0.2, P < 0.05). Furthermore, superfusion of L-arginine (100 microM) caused an increase in capillary Lp (Ltestp/Lcontrolp = 2.4 +/- 0.9, P < 0.05), whereas D-arginine had no effect on Lp (Ltestp/Lcontrolp = 1.2 +/- 0.3). The results of this study support our hypothesis that inhibition of NO synthase decreases capillary Lp in the intact circulation. In addition, L-arginine increases capillary Lp in our model.(ABSTRACT TRUNCATED AT 250 WORDS)

Smart Tissue Culture: in Situ Monitoring of the Activity of Protease Enzymes Secreted from Live Cells Using Nanostructured Photonic Crystals

Nano Letters ◽  
2009 ◽  
Vol 9 (5) ◽  
pp. 2021-2025 ◽  
Author(s):  
Kristopher A. Kilian ◽  
Leo M. H. Lai ◽  
Astrid Magenau ◽  
Siân Cartland ◽  
Till Böcking ◽  
...  
Keyword(s):  
Tissue Culture ◽  
Photonic Crystals ◽  
In Situ Monitoring ◽  
Live Cells

scholarly journals NewIn SituCapture Quantitative (Real-Time) Reverse Transcription-PCR Method as an Alternative Approach for Determining Inactivation of Tulane Virus

2014 ◽  
Vol 80 (7) ◽  
pp. 2120-2124 ◽  
Author(s):  
Dapeng Wang ◽  
Shuxia Xu ◽  
David Yang ◽  
Glenn M. Young ◽  
Peng Tian
Keyword(s):  
Tissue Culture ◽  
Reverse Transcription ◽  
Viral Genome ◽  
Viral Infectivity ◽  
Qrt Pcr ◽  
Reverse Transcription Pcr ◽  
Alternative Approach ◽  
Tulane Virus ◽  
Pcr Method

ABSTRACTHuman noroviruses (HuNoVs) are the major cause of epidemic nonbacterial gastroenteritis. Although quantitative (real-time) reverse transcription-PCR (qRT-PCR) is widely used for detecting HuNoVs, it only detects the presence of viral RNA and does not indicate viral infectivity. Human blood group antigens (HBGAs) have been identified as receptors/co-receptors for both HuNoVs and Tulane virus (TV) and are crucial for viral infection. We propose that viral infectivity can be evaluated with a molecular assay based on receptor-captured viruses. In this study, we employed TV as an HuNoV surrogate to validate the HBGA-based capture qRT-PCR method against the 50% tissue culture infectious dose (TCID50) method. We employed type B HBGA on an immuno-well module to concentrate TV, followed by amplification of the captured viral genome byin situqRT-PCR. We first demonstrated that thisin situcapture qRT-PCR (ISC-qRT-PCR) method could effectively concentrate and detect TV. We then treated TV under either partial or full inactivation conditions and measured the remaining infectivity by ISC-qRT-PCR and a tissue culture-based amplification method (TCID50). We found that the ISC-qRT-PCR method could be used to evaluate virus inactivation deriving from damage to the capsid and study interactions between the capsid and viral receptor. Heat, chlorine, and ethanol treatment primarily affect the capsid structure, which in turns affects the ability of the capsid to bind to viral receptors. Inactivation of the virus by these methods could be reflected by the ISC-qRT-PCR method and confirmed by TCID50assay. However, the loss of the infectivity caused by damage to the viral genome (such as that from UV irradiation) could not be effectively reflected by this method. Despite this limitation, the ISC-qRT-PCR provides an alternative approach to determine inactivation of Tulane virus. A particular advantage of the ISC-qRT-PCR method is that it is also a faster and easier method to effectively recover and detect the viruses, as there is no need to extract viral RNA or to transfer the captured virus from magnetic beads to PCR tubes for further amplification. Therefore, ISC-qRT-PCR can be easily adapted for use in automated systems for multiple samples.

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