Open Blink

Overview

Open Blink is a low-cost TIRF microscopy with laser combiner providing high-power homogeneous illumination for single-molecule based super-resolution imaging. The control system is fully integrated in micro-manager 2.0 .

Open Blink is inspired greatly by K2 TIRF at Ganzinger lab, the LaserEngine at Ries lab, and an flat-field illumination implementation. We also thank the Open Life Science program for helping with the documentation, and Open Hardware Funding at TU Delft for sponsoring parts of the project.

This site includes hardware components used in Open Blink. For the laser combiner and the focus-lock module 'fgFocus', we developed custom control electronics based on microcontrollers, and corresponding Micro-manager plugins, available in https://github.com/GrussmayerLab/LaserController and https://github.com/GrussmayerLab/fgFocus, respectively.

Design files are available in 4TU repository. To cite the project, please refer to our pre-print.

Schematic

Laser Box

The laser box is contained within a 450*450 mm breadboard with enclosure. All lasers are raised up to 70 mm height with customised aluminum heat sink. In a similar layout to Ries Laser Engine, we used two 638 nm lasers to reach high intensity for red excitation demanded by dSTORM. Model numbers are for Thorlabs unless specified otherwise.

Homogeneous illumination is enabled by mode-scrambling of multimode optical fiber (MMF), facilitated with a vibrational motor in vicinity to the MMF.

All the lasers take 0-5V analog modulation, and the motor takes pulse-width-modulation (PWM). Such control signals are generated by a Raspberry Pi board and digital-analog-converters (DACs).

id	Title	Description	Image	Supplier, Model	Cost in €
1	Diode lasers 405 nm	500 mW, with driver	Diode laser and driver	Lasertack, LAB-405-500	687
2	Diode lasers 488 nm	2000 mW, with driver		Lasertack, LAB-488-1000	3310
3	Diode lasers 638 nm, 2x	2x1000 mW, with drivers. Mounted perpendicular to each other.		Lasertack, LAB-638-1000	866
4	DPSS lasers 561 nm	1000 mW, with analog modulation(0-5V)		CNI laser, MGL-FN-561	11150
5	Dichroics	1" longpass dichroic mirros to combine the laser lines		Thorlabs, DMLP425/505/550/605	708
6	PBS	1" polarisation beam spliter, mounted by KM100PM/M and PM4/M, in order to combine two 638 nm lasers	PBS	Thorlabs, PBS251	315
7	Fiber launcher	A 1/2" 19mm ach. doublet (AC127-019-A) to focus the combined beam into the fiber. Mounted in 1" adjustable lens tube (SM05V10, SM1A6, SM1M15, SM1TC), FC/PC adaptor(SM1FC)	Fiber launcher	Thorlabs	220
8	Multimode fiber	70*70 um square profiled MMF, NA=0.22		Ceram Optec, Customized	350
9	Vibrational motor	Vibrational motor is mounted in vicinity to the MMF to reduce the speckles at the MMF output	Vibrational motor Vibrational motor mounting	Precision microdrives, 304-111, or cheaper alternative at Hackerstore.nl	7

Excitation

Similar to K2 TIRF, we used a motorized 1D stage to conveniently switch between Epi-illumination, HILO, and TIRF. Usage of 150 mm focusing lens allows us to achieve TIRF condition from MMF delivery, and provides a large illuminated field of view.

id	Title	Description	Image	Supplier, Model	Cost in €
1	Fiber adaptor	Couple out the beam at the output of the optical fiber mounted on cage plate (CP33/M)		Thorlabs, SM1FC	30
2	Laser collimator	A 30 mm achromatic doublet to collimate the beam at the output of the optical fiber. Mounted in 1" cage after the fiber adaptor	Laser collimator	Thorlabs, AC254-030-A	85
3	Laser cleanup filter	Filter out the autofluorescence from the MMF		Chroma, ZET 405/488/561/640xv2	575
4	TIRF stage	1D motorised stage scans the laser beam across the back focal plane(BFP) of the objective	TIRF stage assembly	Thorlabs, KMTS25E/M	1500
5	TIRF lens	150 mm achromatic doublets to focus the beam onto the BFP of the objective, mounted on CXY1A		Edmund Optics, 147-643	283
6	Elliptical mirror	Mounted on KCB1E/M and assembled with the TIRF lens on the TIRF stage		Thorlabs, BBE1-E02	70

Microscope Body

We reduced the width of the original miCube, so that it can acoommodate the 150 mm TIRF focusing lens. The stage adaptor plate extends above the top of miCube to hold the enclosure.

id	Title	Description	Image	Supplier, Model	Cost in €
1	Modified MiCube	Adapted from MiCube. Reduced in size to accommodate 150 mm focusing lens	Micube	in-house CNC milled	1700
2	XYZ stage	Nano precision sample stage		Smaract, CLS5252	14000
3	Objective	Oil immersion objective, NA = 1,5. Mounted on the micube with M27 to RMS adaptor (RMSA3)		Olympus, UPLAPO60XOHR	12400
4	Quad-band dichroic	Dichroic mirror under the objective, mounted on DFM1/M and fixed on the micube with screws on the back		Chroma, zt405/488/561/640rpc	575
5	Elliptical mirror	Mounted under the dichroic mirror with KCB1E/M and C4W-CC	QUad-band dichroic and mirror mounted inside of Micube	Thorlabs, BBE1-E02	65
6	Sample holder	Home-made slide holder, mounted onto the stage		In-house CNC milled	\
7	Enclosure	Mounted around the adaptor plate on top of the miCube, with an opening for brightfield illumination		Acrylic plates, customized	\

Detection

The detection features a dual-color scheme.

id	Title	Description	Image	Supplier, Model	Cost in €
1	Tube lens	180 mm tube lens compatible with Olympus objectives		Thorlabs, TTL180-A	686
2	Notch filter	Filter out the excitation laser beam		Chroma, ZET405/488/561/640m-TRFv2	1000
3	Slit	Semi rectangular slit adjustable from two directions, cropping the image to fit in a single channel on camera.	Slit	VA100/M	292
4	4f lenses	Three 150 mm achromatic doublet mounted on LMR1/M. One after the tube lens, and one for each color channel. The lens in the red channel is mounted on a small 1D dovetail stage (DT12/M) to co-align both channels.	Two channle 4F system	Edmund Optics, 67-334	342
5	Dichroic mirror	Long-pass dichroic splits image into red and green channels, glued on compact KMS/M with picodent Twinsil		Chroma, ZT640rdc-UF3	450
6	Square mirror	Reflects the green channel with maxium surface, mounted on compact KMS/M		Thorlabs, BBSQ1-E02	74
7	Emission filters	Band-pass filters in the red and green channels respectively		AHF, ET685/70, ET595/50	650
8	Camera	sCMOS camera		Teledyne Prime BSI Express	10800

fgFocus

fgFocus is adapted from pgFocus and qgFocus, and made compatible with micro-manager.

×

This is a sample text to enlarge the div window