Leica SP8 User Manual

Leica SP8 Upright confocal microscope 14, 07, 2020
• System description:
• DMi6, Upright microscope: BF, Fluorescence (blue, green, red)
• BF TL Detector (transmitted light image)
• Scanning stage with z-Galvo and Navigator (overviews, selection of regions of
interest…)
• Scanner with Scan-field Rotation
• 2 PMT detectors (photo multipliers)
• 1 Hybrid Detector normal (more sensitive than PMT and no detector noise nor
background)
• 1 Hybrid Detector SMD (fast, for FLIM, FALCON)
• Laser lines: 405, White Light Laser (470-670nm, up to 8 laser lines simultaneously)
• FALCON (FAst Lifetme CONtrast) including Pulse Picker for WLL
• LIGHTNING (Deconvolution) for Superresolution and super sensitive imaging
• Phasor plot for direct examination of lifetime data
• Objectives:
• Objective HC PL FLUOTAR 5x/0.15 Dry , WD=12mm (overview) (506224)
• Objective HC FLUOTAR L 16x/0.60 IMM CORR VISIR, WD=2.5mm (Correction collar for
use with 0.0 - 0.17 mm cover glass) + BABB cleared samples (Oil, water, glycerol, silicon
oil)- (506533)
• Objective HC FLUOTAR L 25x/0.95 Water-VISIR, WD=2.5 mm, wide angle (41°) (506374)
• Objective PL APO 20x/0.75 IMM CORR VISIR, WD 0.67 mm (Oil, glycerol)- (506343)
• System Start Up
Turn on heating Unit if needed
• Switch on ‘PC-microscope’ button (1). This gives power to the computer and
microscope controller
• Switch on ‘scanner power’ (2)- This enables the scanning head
• Switch on ‘Laser power’ (3)
• Turn the ‘Laser ignition key’ (4) in position ‘on’- enables Laser shutters (-> Only an
interlock)
• Switch on fluorescent lamp(5) for visual examination
• TL/LED manual switch is at the back of the mic (if you don’t see the TL light in the
eyepiece, it might be the switch)

Leica SP8 Specifications

General

Type	Confocal Microscope
Software	LAS X
Scanning Speed	Up to 428 frames per second
Dimensions	Varies depending on configuration
Weight	Varies depending on configuration
Imaging Modes	Confocal
Laser Lines	405 nm, 488 nm, 514 nm, 561 nm, 594 nm, 633 nm
Laser Type	Diode, Argon, DPSS
Scanning Method	Galvanometric
Detectors	Hybrid Detectors
Spectral Detection Range	400 nm - 800 nm
Objectives	10x, 63x oil immersion

Summary

System Description and Objectives

System Start Up Procedure

Initial Power On Sequence

Steps for powering on the microscope, scanner, lasers, and auxiliary components.

LASX Software Configuration

Selecting System Configuration

Laser and Pulse Picker Settings

Instructions for turning on lasers and setting the pulse picker frequency in LASX.

Data Organization and Acquisition

Confocal Image Acquisition Setup

Steps for selecting detectors (PMT/HyD) and acquisition modes in LASX.

Pre-acquisition Checks

Important checks before starting acquisition, like Autosave and HyD detector cooling.

Acquisition Parameter Tuning

Laser and Detector Configuration

System tuning for laser lines, detectors, gain, and emission spectra.

Confocal Image Optimization

Scan Settings

Adjusting format, speed, zoom, and pixel size for optimal image sampling.

Pinhole and SNR Adjustment

Advanced Acquisition Techniques

Gated Detection

Sequential Scan Setup

Configuring Line, Frame, or Stack sequential scans in the acquisition mode.

Sequential Scan and Z-Stack

Z-Stack Setup Procedure

Instructions for setting up Z-stacks using the joystick and 'Z position' knob.

Z-Stack Configuration

Z-Stack Parameters

Selecting Z-galvo or z-Wide, setting start/end points, and Z-step size.

Time Series and Tile Scan

Time-Lapse Settings

Lightning Deconvolution

Choosing Lightning Mode

Lightning Slider Function

Lightning Deconvolution Settings

Deconvolution Strategy Selection

Factory Deconvolution Settings

Laser Safety Instructions

General Laser Safety Rules

Mandatory safety procedures, including warning lights, beam path, and protective coverings.

Personal Protective Equipment

Guidelines for using laser safety goggles and avoiding reflective objects.

User Responsibilities