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3-DIMENSIONAL SCANNING ANALYTICAL LASER MICROSCOPE

Objective

THE POTENTIAL IMPACT CONCERNING DATA CAPTURE AND UPDATING OF DATA BANKS FOR BIOTECHNOLOGICAL/BIOMEDICAL ADVANCEMENT IS EXCEPTIONALLY LARGE. A LASER MICROSCOPE WITH A RESOLUTION BELOW 100 NM AND THREE-DIMENTIONAL DISPLAY COULD HAVE GREAT IMPACT ON BIOTECHNOLOGY AND FIND WIDESPREAD APPLICATION. SPECIFIC ADVANTAGES OVER ELECTRONIC MICROSCOPY INCLUDE : NO NEED FOR VACUUM, THE USE OF LIVE SPECIMENS IN THEIR NATURAL ENVIRONMENTS; AND THE ABILITY TO STUDY SPECIMEN DEVELOPMENT DURING TIME. THERE IS A POTENTIALLY LARGE MARKET IN WHICH EUROPE WOULD TAKE THE LEAD AND THUS EMPLOYMENT TO BE CREATED IN THE EVENT OF SUCCESSFUL DEVELOPMENT.
Starting with the instrument in Amsterdam, the program had the objectives of developing suitable 3-dimensional image processing and routine analyses for extracting the maximum amount of information from the confocal data, and 3D superresolution approaches based on a combination of physical and mathematical techniques available. These efforts were supported on the optical and technical side by the development of special objects, and test specimen and wavefront measurements, while biological test specimens and preparation techniques were supplied from Utrecht.

Using a prototype laser confocal microscope existing in Amsterdam, research was carried out in order to expand the scope and range of application of this type of microscopy. The addition of novel imaging modes, superresolution techniques and dedicated 3-dimensional image processing would make this technique a powerful analytical tool for biology.

A double detector system was added to the Amsterdam instrument for multiple label detection in biological systems. Use of variable microdiaphragms in illumination and detection provided optical probe control. Data inversion methods were developed which use a Fourier transform and Tikhonov regularization. These methods were applied to images of biological samples and increased confocal axial resolution was achieved. A novel method was conceived and tested experimentally at low NA for achieving confocal superresolution by holographic processing of the signals in the detection plane. Special optics were developed. A dedicated 3-dimensional image processing and analysis package was developed for the processing of confocal images.
THE PROJECT IS A MULTI-NATIONAL COLLABORATION BETWEEN A NUMBER OF LEADING RESEARCH AND INDUSTRIAL ESTABLISHMENTS WHOSE LONG-TERM OBJECTIVE IS TO ADVANCE SIGNIFICANTLY THREE-DIMENTIONAL AND SUPER-RESOLUTION SCANNING LASER MICROSCOPY. THE BACKGROUND IS THE EXISTING PROTOTYPE CONSTRUCTION OF A LASER-BASED CONFOCAL SCANNING OPTICAL MICROSCOPE AT THE CONTRACTORS LABORATORY. THE SPECIFIC AIM OF THE PROJECT IS TO INTRODUCE A WIDE RANGE OF NEW TECHNIQUES INTO SCANNING LASER MICROSCOPY, BASED ON THE ABOVE MENTIONED SYSTEM. SIX OTHER GROUPS WILL PARTICIPATE IN THE PROJECT (AS SUB-CONTRACTOR) THUS AIMING AT THE ENHANCEMENT OF THE THREE-DIMENSIONAL AND SUPER-RESOLVING (BELOW 100 NM) CAPABILITIES.

Funding Scheme

CSC - Cost-sharing contracts

Coordinator

UNIVERSITEIT VAN AMSTERDAM
Address
Maagdenhuis Spui 21
Amsterdam
Netherlands

Participants (5)

LEITZ
Germany
Address

Wetzlar
National Physical Laboratory (NPL)
United Kingdom
Address
Queen's Road
TW11 0LW Teddington
Public Health Laboratory Service (PHLS)
United Kingdom
Address
61 Colindale Avenue
NW9 5HT London
UNIVERSITÀ DEGLI STUDI DI GENOVA
Italy
University of Utrecht
Netherlands
Address

3508 Utrecht