Single Molecule Detection | PL1

Overview of PL1

A time-resolved confocal microscope, the PL1 is designed primarily for material sciences research requiring the ultimate sensitivity in FLIM acquisition of large area samples. The sample, with dimensions up to 100 x 100 mm for the upright microscope and up to 120 x 75 mm for the inverted type, is placed on the high-precision, computer-controlled, XY stage that travels from pixel to pixel featuring a 22 nm resolution.

The excitation source is a laser diode, a pulsed laser, or a multiphoton laser. The fluorescence is collected by one detector covering the range 350 – 1050 nm; additional detectors can be added including a spectrograph for pixel spectral acquisition.

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Key Features of PL1

FLIM of Large Areas

Upright Microscope:
100 mm x 100 mm
Inverted Microscope:
120 mm x 75 mm

Lifetime Measurements

From 100 ps to 100 ms

Modularity

A selection of laser wavelengths, detectors, number of detection channels, & microscopes.

The Data is Clear!

Quantum dots fluorescence map — Phasor plots separate different lifetime species of quantum dots coated on the substrate directly from the raw data. Excitation was 470 nm, emission: 499 - 632 nm (fluorescence), 475/35 nm (reflection), scanning area 17.5 µm x 17.5 µm.
*(Courtesy of Wenjie Liu and Dr. Yaowu Hu; Purdue University; West Lafayette, IN; USA)*

Quantum dots reflection map — Phasor plots separate different lifetime species of quantum dots coated on the substrate directly from the raw data. Excitation was 470 nm, emission: 499 - 632 nm (fluorescence), 475/35 nm (reflection), scanning area 17.5 µm x 17.5 µm.
*(Courtesy of Wenjie Liu and Dr. Yaowu Hu; Purdue University; West Lafayette, IN; USA)*

Perovskite confocal lifetime imaging for both photoluminescence intensity and lifetime at the high spatial resolution. ISS VistaVision provides both the minimization fitting algorithm and the phasor plots for lifetime analysis; for example, using the multi-image phasor analysis routine, the lifetime changes in different Perovskite samples can be immediately identified by the phasor plots of their raw data sets (no fitting required), making the lifetime analysis much easier and robust. Excitation was 488 nm; emission through a 635 nm long pass filter; scanning area 20 µm x 20 µm.
*(Courtesy of Dr. Xiaodan Zhang; Nan Kai University, Tianjin, P.R. China)*

Confocal lifetime imaging for nanoparticle upconversion — Upconversion nanoparticle confocal lifetime imaging for both photoluminescence intensity and lifetime at the high spatial resolution. Excitation: 980 nm, Emission: 509 - 552 nm, scanning area 20 µm x 20 µm
*(Courtesy of Dr. Yan Guan, Peking University, Beijing; P.R. China)*

Product Specifications for PL1

Microscope and Coupling

Frame format: Upright or Inverted research microscope
Magnification: 10X and 60X, oil immersion objective (standard); optional: from 2X to 100X
Spatial Resolution: diffraction limited
Eye Observation: bright field by 10X eyepiece with diopter adjustment, field of view: 22 mm
Imaging Modes:
- Transmission mode: HAL Köhler illumination for bright field imaging by a CMOS camera with options for phase contract and DIC
- Confocal Photoluminescence imaging: laser illumination, single point or scan

XYZ Stage Scan

Closed-loop DC servo control

XY axis range of travel: 100 mm x 100 mm (upright), 120 mm x 75 mm (inverted)
XY axis: Resolution = 22 nm, max velocity = 7 mm/s, RMS repeatability
Z axis: Resolution = 50 nm, maximum velocity = 0.6 mm/s, repeatability = 100 nm

Laser Sources

CW or pulsed diode laser, repetition rate up to 80 MHz (tunable by software)
The laser launcher can accommodate up to 3 lasers, ranging from 375 nm to 980 nm
Each laser has its own intensity control and shutter (operated by software)

Data Acquisition Unit FastFLIM

Lifetime measurement range: from 100 ps to 100 ms
Data Acquisition Mode: Photon mode, Time-tagged mode, Time-resolved time-tagged mode (TTTR)
Dead Time: 3.125 ns, Up to 60 x 10⁶ counts/second
Computer Connection: USB

Detectors

Dark Counts < 100/sec; TTS < 350 ps;
Wavelength Range: 350 - 1050 nm;
Quantum Efficiency > 70% at 700 nm

Scan Modes

X, XY, XZ, XYZ, t, Xt, XYt, XZt, XYZt
Image Format Other than the proprietary file formats which contain the imaging parameters information, VistaVision also supports exporting the acquired data in various formats including JPEG, TIFF, PNG, and AVI.
Image Processing and Analysis Visualization by various look–up tables, contrasting, thresholding, smoothing, filtering, scaling, statistical analysis by histogram or online profiling.

Lifetime Data Analysis Routines

Non-linear least square constrained deconvolution fitting routines based on the Marquardt-Levenberg minimization algorithm in both time and frequency domains
Model-free phasor plots approach for instant and unbiased results

Software

VistaVision

Computer & Monitor

High-performance Processor, 32 GB RAM, Windows 10, 64-bit
32" monitor, 2556 x 1440 resolution

Power Requirements

Universal power input: 110 - 240 V, 50/60 Hz, 100 VAC

Measurements for PL1

Intensity and Lifetime Imaging

Fluorescence Fluctuations Spectroscopy (FFS)

Single Molecule FRET Bursts Analysis

Configuration for PL1

Photoluminescence Microscope Configuration

Product Software for PL1

VistaVision

VistaVision is a complete software package for confocal microscopy applications including instrument control, data acquisition and data processing. Easy to use, the software has been developed in modular components that can be activated when a specific instrument configuration is selected. The modules include:

FLIM/PLIM Imaging
FFS
smFRET
Particle Tracking

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Product Resources

Application Notes

Technical Notes

PL1Single Molecule DetectionCompact Instrument