Increasing the sensitivity and resolution of single plane light sheet microscopy with multi-pulse pumping and time-gated detection (Conference Presentation)

Joseph D. Kimball; Zhangatay Nurekeyev; Jose Chavez; Luca Ceresa; Sangram Raut; Ignacy Gryczynski; Zygmunt Gryczynski

doi:10.1117/12.2514298

4 March 2019 Increasing the sensitivity and resolution of single plane light sheet microscopy with multi-pulse pumping and time-gated detection (Conference Presentation)

Joseph D. Kimball, Zhangatay Nurekeyev, Jose Chavez, Luca Ceresa, Sangram Raut, Ignacy Gryczynski, Zygmunt Gryczynski

Author Affiliations +

Proceedings Volume 10884, Single Molecule Spectroscopy and Superresolution Imaging XII; 1088405 (2019) https://doi.org/10.1117/12.2514298
Event: SPIE BiOS, 2019, San Francisco, California, United States

Abstract

Optical microscopes have proven their use as a powerful tool for studying a variety of biological samples. In spite of many successes, there are still numerous obstacles limiting practical applications. Most limiting are the inherent background of physiological samples, photobleaching, and phototoxicity. To allow studies of long lasting processes such as drag delivery, three-dimensional cellular structures, embryogenesis, we have combined a technique called Single Plane Illumination Microscopy (SPIM) with Multi-Pulse Pumping with Time-Gated Detection (MPP-TGD) in order to enhance the signal relative to background. This new method allows for a decrease in light exposure times and improves image quality. This combination allows a new outlook into a variety of important, long-lasting biological processes at a level of detection previously unattainable. Multi-pulse pumping is a burst of excitation pulses instead of a single pulse which enhances the excited state population of a long-lived label. This label is chosen so that its lifetime is at least 5 times longer than that of typical autofluorescence. The pulse separation within the burst is chosen so that it is at least 5 times shorter than the lifetime of the label. In this case only the population of the fluorescent label is increased and the background remains the same. By subtracting the image acquired with the burst from an image with a single pulse, we were able to increase the signal-to-background ratio of about 100 fold.

Conference Presentation

Citation Download Citation

Joseph D. Kimball, Zhangatay Nurekeyev, Jose Chavez, Luca Ceresa, Sangram Raut, Ignacy Gryczynski, and Zygmunt Gryczynski "Increasing the sensitivity and resolution of single plane light sheet microscopy with multi-pulse pumping and time-gated detection (Conference Presentation)", Proc. SPIE 10884, Single Molecule Spectroscopy and Superresolution Imaging XII, 1088405 (4 March 2019); https://doi.org/10.1117/12.2514298

ACCESS THE FULL ARTICLE

INSTITUTIONAL
Select your institution to access the SPIE Digital Library.

SELECT YOUR INSTITUTION

PERSONAL
Sign in with your SPIE account to access your personal subscriptions or to use specific features such as save to my library, sign up for alerts, save searches, etc.

PERSONAL SIGN IN

No SPIE Account? Create one

PURCHASE THIS CONTENT

SUBSCRIBE TO DIGITAL LIBRARY

50 downloads per 1-year subscription

Members: $195

Non-members: $335 ADD TO CART

25 downloads per 1 - year subscription

Members: $145

Non-members: $250 ADD TO CART

PURCHASE SINGLE ARTICLE

Includes PDF, HTML & Video, when available

Members: $17.00

Non-members: $21.00 ADD TO CART

PROCEEDINGS
PRESENTATION

WATCH
PRESENTATION SAVE TO MY LIBRARY

GET CITATION

KEYWORDS

Microscopy

Signal processing

Fluorescent markers

Image quality

Optical microscopes

Signal detection

Keywords/Phrases

Search In:

Publication Years