Multiphoton Microscopy
Femtosecond excitation, dispersion compensation, scanning, and detection for two-photon, three-photon, and deep-tissue imaging.
Step 1 — Define your goal
What are you trying to achieve?
Pick the experiment / project closest to yours. We'll route you to the right system architecture and BOM.
Step 2 — Confirm the problem
Common project challenges
If any of these sound familiar, you're in the right place. WaveQuanta engineers have seen — and solved — every one of them.
Excitation wavelength selection
GCaMP @ 920 nm, RFP @ 1040 nm, deep brain @ 1300 nm — pick the matrix.
Do I need an OPA?
If you want wavelength tunability or 3P at unusual wavelengths, yes.
Dispersion compensation
GDD from objective + scanner can stretch your pulse from 100 fs to > 300 fs at the sample. Need pre-comp.
Pulse width at the sample
Verify with autocorrelator at the focal plane — what comes out of the laser ≠ what's at the sample.
Scanning architecture
Galvo / resonant / AOD — depends on frame rate and FOV target.
Detection optics + filters
PMT / GaAsP / hybrid — paired with emission filters for each fluorophore.
Photodamage management
Average power and pulse density at the sample — avoid bleaching or thermal damage.
Scope integration mechanics
Beam steering into a commercial scope (Olympus / Nikon / Bruker / Sutter).
Step 3 — Understand the system
Typical system architecture
Most projects in this area follow a similar signal flow. Your specific architecture depends on resolution, throughput, and form-factor targets.
Ti:Sapph at 800 nm or Yb at 1030 nm — your standard 2P excitation source.
OPA gives access to wavelengths the laser doesn't produce (1300, 1700 nm for 3P).
Pre-compensate dispersion of the objective + scanner. Fs pulses must arrive sharp at the sample.
Power control, beam expansion, polarization for the scope's input aperture.
Galvo or resonant — resonant gives kHz frame rate, galvo gives flexibility.
Step 4 — Pick the modules
Recommended system modules
These are the building blocks. Each module is a category of products — pick the right brand and grade for your project stage below.
Femtosecond Laser Source
Ti:Sapph at 800 nm or Yb at 1030 nm — your standard 2P excitation source.
- Ti:Sapph 690–1020 nm tunable
- Yb 1030 / 1040 nm fixed
- Sub-100 fs pulse width
- MHz repetition rate
Tunable OPA / Multi-Color
OPA gives access to wavelengths the laser doesn't produce (1300, 1700 nm for 3P).
- Yi-Laser AURORA OPA
- Signal 1.2–2 µm
- Idler 2–4 µm
- Non-collinear option for 3P
Pulse Compression / GDD Comp.
Pre-compensate dispersion of the objective + scanner. Fs pulses must arrive sharp at the sample.
- Prism pair compressor
- Chirped mirror set
- Dazzler for high-precision GDD
Pre-Scope Beamline
Power control, beam expansion, polarization for the scope's input aperture.
- λ/2 + PBS attenuator
- Beam expander to scope aperture
- Flat-top polarization
Scanning Optics
Galvo or resonant — resonant gives kHz frame rate, galvo gives flexibility.
- Galvo X-Y for benchtop
- Resonant for high frame rate
- AOD for ultra-fast random access
Objective + Sample
High-NA water-dipping objective with NIR / IR transmission.
- Olympus XLPLN25XWMP for 2P
- Nikon CFI Plan Apo IR 60×
- Leica HCX IRAPO L for 3P
Fluorescence Detection
PMT / GaAsP / hybrid for low-light fluorescence; pulse-counting for sparse signal.
- Hamamatsu GaAsP PMT
- Hybrid PMT for fast counting
- Multi-channel filter wheel
Filter / Dichroic Path
Emission separation between channels; chromatic correction at high NA.
- Multiband dichroic stack
- Emission filter wheel
- Chromatic correction lens
Step 5 — Match your project stage
Choose your project stage
Same modules, three configurations sized for where your project is today. Move up the tiers as you progress from research to validation to OEM.
Research Starter
PhD lab / single scope
Add fs excitation to an existing scope. Validates your sample-fluorophore-power workflow.
- Compact Yb / Ti:Sapph fs laser
- Stock pre-comp prism pair
- Galvo + photodiode
- Manual filter wheel
- Integration kit for common scopes
BOM tier: $60k – $180k
Engineering Validation
Imaging core / 2P + 3P combo
Locked-spec multiphoton excitation engine. Multi-wavelength + tunable OPA + GDD comp + fast scanner.
- Yi-Laser CARMEL / SPECTRA-X 2P fs laser
- AURORA OPA for 3P / multi-color
- Dazzler / chirped mirror GDD comp
- Resonant / galvo scanner combo
- GaAsP PMT array
- Full integration & alignment
BOM tier: $250k – $700k
OEM Production
Microscope OEM · turnkey scope
Productized excitation engine for a commercial multiphoton microscope. Locked BOM, factory alignment, supply contract.
- Locked-spec fs laser engine
- Integrated OPA module
- Factory pre-aligned subassembly
- Field-replaceable filter cubes
- Full QA documentation
- 5+ year supply
BOM tier: $700k+ · NRE + per-unit
Step 6 — Run the numbers
Recommended calculators
Sanity-check your design before talking to an engineer.
Step 7 — Configure the system
Configure your setup with our engineering tools
Two ways to go from "this is what I want to do" to "this is the BOM I need".
Open Multiphoton Optical Path VL
Configure laser, OPA, GDD comp, scanner, and detector. Verify pulse width at the sample and dispersion budget.
Launch Virtual LabAsk AI to spec my multiphoton excitation
Describe samples, fluorophores, frame rate, and target imaging depth. AI Concierge proposes laser, OPA, GDD comp, and detection.
Open AI ConciergeStep 8 — Anchor SKUs
Featured products for this application
Common picks for this application area. Your final BOM will pull from a wider catalog and be sized to your project tier.
Yi-Laser
HELIOS Femtosecond Laser
Yi-Laser
AURORA OPA — High Energy
Yi-Laser
HYPERION-G MPC Pulse Compressor
Fastlite
Dazzler Pulse Shaper
Princeton Instruments
Princeton sCMOS Camera
Ultrafast Innovations
Chirped Mirror — GDD Compensation
LBTek
Longpass Dichroic Mirror DM20-638LP
LBTek
Longpass Dichroic Mirror
Step 9 — Common questions
Frequently asked questions
Quick answers to the questions our application engineers hear most often.
Ti:Sapph or Yb fs — which for multiphoton?
Ti:Sapph (690–1020 nm tunable): most flexible for 2P, gold standard for GCaMP / GFP. Yb (1030–1060 nm): cheaper, more reliable, fixed wavelength. Best for RFP / mCherry imaging. Most modern labs run Yb for 2P + OPA for tunability.
How important is dispersion compensation?
Critical. A 100 fs pulse passing a high-NA water objective + scanner stretches to 300+ fs without pre-compensation. That kills your 2P signal (which scales as 1/τ²). Always include a chirped mirror set or Dazzler.
Do I need an OPA for 2P?
If your fluorophore has good 2P cross-section at your laser wavelength: no. If you want to multiplex 2 fluorophores or do 3P imaging at >1300 nm: yes.
How deep can 3P see in mouse brain?
State-of-the-art: 1.2 mm cortical depth in mouse, including white matter. Requires sub-100 fs pulse at 1300 / 1700 nm with 1+ µJ pulse energy.
Pulse rate — MHz or kHz?
Most 2P labs use 80 MHz. For high-power 3P or deep imaging, drop to 1–4 MHz to keep average power manageable. Pulse picker can convert MHz to lower rep rate.
Resonant vs galvo scanner?
Galvo: arbitrary scan paths, slower (Hz–kHz frames). Resonant: 30+ Hz at 512×512, but fixed sinusoidal trajectory. Most labs run resonant + galvo combo.
Photodamage — how to avoid?
Keep average power at sample < 30 mW for cortical 2P, < 100 mW for 3P. Use minimum fluence for SNR — don't blast the sample. Pulse-train modulation avoids thermal accumulation.
Can I integrate this with existing Olympus / Nikon / Bruker scopes?
Yes — WaveQuanta supplies integration kits with periscope, beam expander, and motorized power control. For Bruker / Sutter Ultima / Thorlabs Bergamo: drop-in compatible.
Step 10 — Engineering Review
Application Engineering Review
Tell us your application, current setup, and project context. A WaveQuanta application engineer will return initial recommendations within 1 business day.
- 1 Application
- 2 Current setup
- 3 Project & purchase