Contamination of surfaces prior to optical coating by in-situ total internal reflection microscopy

1994 ◽  
Author(s):  
David W. Reicher ◽  
Kenneth C. Jungling ◽  
Charles K. Carniglia
Keyword(s):  
Total Internal Reflection ◽  
Internal Reflection ◽  
Optical Coating ◽  
Total Internal Reflection Microscopy

scholarly journals In Situ Fluorescent Protein Imaging with Metal Film-Enhanced Total Internal Reflection Microscopy

2006 ◽  
Vol 90 (12) ◽  
pp. 4662-4671 ◽  
Author(s):  
Thomas P. Burghardt ◽  
Jon E. Charlesworth ◽  
Miriam F. Halstead ◽  
James E. Tarara ◽  
Katalin Ajtai
Keyword(s):  
Total Internal Reflection ◽  
Metal Film ◽  
Fluorescent Protein ◽  
Internal Reflection ◽  
Protein Imaging ◽  
Total Internal Reflection Microscopy

Measurement of Ca2+ signaling dynamics in exocrine cells with total internal reflection microscopy

2006 ◽  
Vol 291 (1) ◽  
pp. G146-G155 ◽  
Author(s):  
Jong Hak Won ◽  
David I. Yule
Keyword(s):  
Plasma Membrane ◽  
Total Internal Reflection ◽  
Acinar Cells ◽  
Internal Reflection ◽  
Pancreatic Acinar Cells ◽  
Wide Field ◽  
Exocrine Cells ◽  
Parotid Acinar Cells ◽  
Signaling Dynamics ◽  
Total Internal Reflection Microscopy

In nonexcitable cells, such as exocrine cells from the pancreas and salivary glands, agonist-stimulated Ca2+ signals consist of both Ca2+ release and Ca2+ influx. We have investigated the contribution of these processes to membrane-localized Ca2+ signals in pancreatic and parotid acinar cells using total internal reflection fluorescence (TIRF) microscopy (TIRFM). This technique allows imaging with unsurpassed resolution in a limited zone at the interface of the plasma membrane and the coverslip. In TIRFM mode, physiological agonist stimulation resulted in Ca2+ oscillations in both pancreas and parotid with qualitatively similar characteristics to those reported using conventional wide-field microscopy (WFM). Because local Ca2+ release in the TIRF zone would be expected to saturate the Ca2+ indicator (Fluo-4), these data suggest that Ca2+ release is occurring some distance from the area subjected to the measurement. When acini were stimulated with supermaximal concentrations of agonists, an initial peak, largely due to Ca2+ release, followed by a substantial, maintained plateau phase indicative of Ca2+ entry, was observed. The contribution of Ca2+ influx and Ca2+ release in isolation to these near-plasma membrane Ca2+ signals was investigated by using a Ca2+ readmission protocol. In the absence of extracellular Ca2+, the profile and magnitude of the initial Ca2+ release following stimulation with maximal concentrations of agonist or after SERCA pump inhibition were similar to those obtained with WFM in both pancreas and parotid acini. In contrast, when Ca2+ influx was isolated by subsequent Ca2+ readmission, the Ca2+ signals evoked were more robust than those measured with WFM. Furthermore, in parotid acinar cells, Ca2+ readdition often resulted in the apparent saturation of Fluo-4 but not of the low-affinity dye Fluo-4-FF. Interestingly, Ca2+ influx as measured by this protocol in parotid acinar cells was substantially greater than that initiated in pancreatic acinar cells. Indeed, robust Ca2+ influx was observed in parotid acinar cells even at low physiological concentrations of agonist. These data indicate that TIRFM is a useful tool to monitor agonist-stimulated near-membrane Ca2+ signals mediated by Ca2+ influx in exocrine acinar cells. In addition, TIRFM reveals that the extent of Ca2+ influx in parotid acinar cells is greater than pancreatic acinar cells when compared using identical methodologies.

Attenuated Total Internal Reflection Infrared Microspectroscopy For The Study Of Trace Contaminants In Aqueous Solutions.

1999 ◽  
Vol 5 (S2) ◽  
pp. 66-67
Author(s):  
Andre’ J. Sommer ◽  
Mark Hardgrove
Keyword(s):  
Infrared Spectroscopy ◽  
Total Internal Reflection ◽  
Molecular Spectroscopy ◽  
Internal Reflection ◽  
Small Samples ◽  
Attenuated Total Internal Reflection ◽  
Infrared Microspectroscopy ◽  
Transmission Measurements ◽  
Major Emphasis

Over the past several years many developments have taken place in the field of molecular spectroscopy. For Raman spectroscopy many of the improvements have arisen from technological innovations that include diode-based lasers, holographic notch filters and charged coupled detectors. In contrast, a majority of the developments in infrared spectroscopy have been in the area of new sampling accessories. A major emphasis has been placed on attenuated total internal reflection (ATR) accessories. The devices are allowing infrared spectroscopy to be employed in process control environments and quality control laboratories where the method is not only robust but has the advantages of limited sample preparation and/or in situ analysis.In the realm of microspectroscopy, ATR accessories have the added advantages of providing better spatial resolution, equal to or higher S/N for equivalent sample size compared to transmission measurements and most importantly the ability to collect spectra of small samples without the adverse effect of diffraction. One accessory which was developed several years ago is known as the Split-Pea.

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