Test page

Category
Description	Samara scratchpad

What's Inside Your Miniscope V4 Miniscope V4 Guide 2026-07-20 1.0 Aharoni Lab A plain-language walkthrough of what each part of an assembled Miniscope V4 does and why, written for researchers imaging animals — not hardware developers. Hardware Optics Reference Beginner End users

The basic idea

Your Miniscope is a fluorescence microscope shrunk down to fit on an animal's head. It shines light of one color into the tissue to excite a fluorescent indicator that the neurons express (commonly a GCaMP-family calcium indicator), then collects the different, much dimmer color of light that comes back and focuses it onto a tiny camera sensor. Almost everything else described below exists for one reason: to keep those two colors of light — the bright light going in, and the faint signal coming back — from mixing.

2.6 g

Mass

22 mm

Height

5 × 5 mm

Baseplate

≥ 1 mm

Field of view

±200 µm

Focus range

0.3 mm coax

Cable

The optical path

Fluorescence microscopes separate excitation and emission light using three parts working together, sometimes called a filter cube: an excitation filter, a dichroic mirror, and an emission filter. The Miniscope's whole optical stack is a miniaturized version of that same idea, folded into one small barrel.

Excitation path: Blue LED → collector lens → excitation filter → dichroic mirror → tissue

Emission path: Tissue fluorescence → objective lens → dichroic mirror → emission filter → camera sensor

Excitation LED & filter — the light source

In general: LEDs put out a broad smear of wavelengths, not one clean color. Left alone, that spread would bleed into the same range as the faint signal you're trying to collect. An excitation filter is a bandpass filter — it only lets a narrow, specific slice of color through, cleaning up the LED's output before it ever reaches the tissue.

In this scope, the LED is blue (~450 nm), and its light first passes through a small half-ball lens that collects and focuses it, then through the excitation filter, which narrows it to a clean, consistent band around ~470 nm. That's the color actually reaching your tissue on every trial — the same wavelength that drives common green calcium indicators.

Dichroic mirror — the beam-splitter

In general: A dichroic mirror looks like a mirror but is picky about color — it reflects light on one side of a wavelength cutoff and lets longer wavelengths pass straight through it. Mounted at 45°, it lets the same objective lens both deliver excitation light down to the tissue and collect the return signal back up, without a separate light path for each direction.

Here, the dichroic reflects the blue excitation light downward through the objective and into the tissue, while letting the longer-wavelength green fluorescence coming back from the tissue pass straight through it, up toward the camera.

Emission filter — the final cleanup

In general: No dichroic is perfect — a little reflected excitation light always leaks through in the same direction as the real signal. An emission filter is a second, stricter color check placed right before the camera, so only genuine fluorescence reaches the sensor rather than stray excitation light.

This scope's emission filter passes a band around ~525 nm (green) — matching where common green indicators emit — and blocks the blue excitation light that would otherwise wash out or contaminate your recording.

Relay & objective lenses — forming the image

In general: Ordinary lenses focus different colors of light slightly differently — a well-known optical flaw called chromatic aberration. Achromatic lenses are designed to bring multiple wavelengths to the same focus, so the excitation path and emission path both stay sharp through the same optics.

A fixed relay lens carries the image from the dichroic down toward the tip; the two lenses at the very tip are the swappable "objective" pair that actually faces the tissue. Which pair is installed sets your working distance (how far the lens sits from the tissue) and field of view (how much tissue you can see at once) — see the configuration table below.

Electrowetting (liquid) focus lens — electronic focus

In general: Most cameras focus by physically sliding a lens back and forth. This scope instead uses a sealed chamber of two liquids that don't mix — applying a small voltage reshapes the interface between them, which changes the lens's focal length. No motors, no moving parts, nothing to jostle out of alignment while an animal is moving around.

This is what lets you adjust focus from the DAQ software (±200 µm of travel) after the scope is already mounted and the animal is behaving — useful for fine-tuning focus session to session without touching the hardware.

Objective lens configurations

Tip: If you're not sure which configuration your scope shipped with, check your order details — the pair of lenses at the tip is what's swapped between configurations, and it's the main thing that determines how much tissue you can see and how far the lens needs to sit from your cranial window.

Config	Working distance	Field of view
1 (default)	0.7 mm	1.0 × 1.0 mm
2	1.0 mm	1.1 × 1.2 mm
3	2.0 mm	1.3 × 1.4 mm

File:EWL.png

Same field of view, three focus settings — only the electrowetting lens voltage changed

Electronics — what's actually running inside the scope

All of the scope's electronics live on one small board folded into four sections. You'll never interact with these directly, but knowing roughly what each does helps make sense of what the DAQ software is actually controlling.

File:PCB Labeled.PNG

The same board, flattened out before it's folded into the scope

Camera: The actual image sensor — a small monochrome camera chip that sits right where the emission light comes to focus. It converts the collected fluorescence into the video frames you see live in the DAQ software.

Power & data: Everything the scope needs travels down the single thin coaxial cable connecting it to the DAQ: incoming power, and outgoing video — packed onto that one cable so the animal only ever carries one lightweight, flexible wire instead of a bundle.

Control & head orientation: A small onboard controller applies the settings you choose in the DAQ software — camera gain, LED brightness, imaging mode — in real time. It also hosts a 9-axis orientation sensor, similar to the one in a phone, which reports the absolute tilt and rotation of the animal's head. You can log this alongside your recording to relate neural activity to head movement and orientation during analysis.

Illumination & focus driver: Supplies current to the excitation LED and voltage to the electrowetting focus lens. Both the LED brightness and the focus setting you adjust in software ultimately come down to this board turning those software values into the actual current or voltage each part receives.

Mounting: baseplate & holder

Baseplate: Cemented to the skull around the cranial window during the scope's first attachment surgery. After that, the scope itself docks onto the baseplate for every subsequent session, returning to the same field of view each time without needing to be re-aligned. 5 × 5 mm footprint.

Stereotaxic holder: A separate 3D-printed fixture used only during that baseplating surgery, to hold the scope perfectly still on a stereotaxic frame while the baseplate is cemented in place. It's not part of the scope you actually record with.

Not covered here

Scope of this guide:

DAQ software & recording settings — see Miniscope V4: DAQ & Recording Settings.
Baseplating surgery — see your lab's surgical SOP for implantation and cementing procedure.
Cleaning up recordings — see the V4 Denoising Notebook if you see faint horizontal banding in your video.

Wavelength numbers above (450 nm, ~470 nm, ~525 nm) describe roughly what part of the spectrum each stage is working with — useful for knowing which fluorescent indicators pair well with this scope. For precise passband numbers, check the specific filter's datasheet.