Okolab H201-NIKON-TI-S-ER User manual

H201-NIKON-TI-S-ER User Manual
August 2019
www.oko-lab.com
Page1
H201-NIKON-TI-S-ER
Compatible with the following XY stages
Compatible with the following Okolab gas modules
•Nikon: Ti-S-E and Ti-S-ER
IST450_rev01
•CO2-UNIT-BL
•CO2-UNIT-BL-CP2
•CO2-O2-UNIT-BL-CP2
•O2-UNIT-BL [1-20]
•CO2-O2-UNIT-BL [0-10;1-18]
•CO2-O2-UNIT-BL [0-20;1-95]
•CO2-O2-UNIT-BL [0-10;0-1]
•2GF-MIXER
•3GF-MIXER-HYPOXIA
•3GF-MIXER-HYPEROXIA

H201-NIKON-TI-S-ER User Manual
August 2019
www.oko-lab.com
Page2
Index
1. Components and dimensions................................................................................................... 3
2. Available Sample Holders....................................................................................................... 3
3. Available Lids ..................................................................................................................... 4
3.1 Sliding lid .......................................................................................................................... 5
3.2 Koehler Lid........................................................................................................................ 6
3.3 Injection Lid ...................................................................................................................... 6
3.4 Laser Interlock Lid............................................................................................................... 7
4. Insertion of the Sample Feedback Temperature Sensor.................................................................. 8
5. Insertion of Sample Holder into Chamber ................................................................................... 8
6. Working with 35 or 60 mm Petri Dish –Spacing Rings and Magnetic Locks ........................................... 9
7. Working with 1x3’’ and 1x2’’ chamber slides - magnetic locks........................................................11
8. Working with MW Plates - Magnetic locks and Chamber riser ..........................................................12
9. Connection of the Gas Supply ................................................................................................13
10. Working with Perfusion ........................................................................................................13
11.Connection of the Chamber with adapter..................................................................................16
12. Connection of the Chamber with XY stage .................................................................................17

H201-NIKON-TI-S-ER User Manual
August 2019
www.oko-lab.com
Page3
1. Components and dimensions
H201-NIKON-TI-S-ER includes the following components:
•Chamber main body. It connects to the adapter with screws.
•Sliding glass lid for easy pipetting.
•Chamber riser. It is a removable frame increasing the height of the chamber from 24 to 30 mm, often re-
quired when using multi-well (MW) plates.
•Adapter. It connects to the Nikon XY Stage TI-S-ER with screws.
Distance between focal plane
and upper side of the glass
lid without the chamber riser
Distance between focal plane
and upper side of the glass
lid with the chamber riser
Chamber top view
Sliding Lid
Chamber Riser
Chamber Base
Sample Holder-Order Separately
Adapter
Figure 1. H201-NIKON-TI-S-ER - Components and Dimensions.
2. Available Sample Holders
The following sample holders are available.
NOTE: Please contact [email protected] if you cannot find the sample holder you are looking for. We are constantly
adding new inserts to the list.
1x35-M
#1 35mm Petri-dish
1xGS-M
#1 1x3in. chamber slide
1xLABTEK-M
#1 Lab-Tek 1in.x2in. chambered cover glass
1xLABTEK-II-M
#1 Lab-Tek II 1in.x2in. chambered cover glass
1x60-M
#1 60mm Petri-dish
1xT25-M
#1 Nunc and Greiner T25 flask
2x35-M
#2 35mm Petri-dish
2xGS-M
#2 1x3in. chamber slides
2xLABTEK-M
#2 Lab-Tek 1in.x2in. chambered cover glass
2xLABTEK-II-M
#2 Lab-Tek II 1in.x2in. chambered cover glass
2x60-M
#2 60mm Petri-dish

H201-NIKON-TI-S-ER User Manual
August 2019
www.oko-lab.com
Page4
4x35-M
#4 35mm Petri-dish
GS35-M
#1 1x3in. chamber slide and #2 35mm Petri-dish
LABTEK-35-M
#1 Lab-Tek 1in.x2in. chambered cover glass and #2 35mm Petri-dish
LABTEK-II-35-M
#1 Lab-Tek II 1in.x2in. chambered cover glass and #2 35mm Petri-dish
LABTEK-60-M
#1 Lab-Tek 1in.x2in. chambered cover glass and #1 60mm Petri-dish
LABTEK-II-60-M
#1 Lab-Tek II 1in.x2in. chambered cover glass and #1 60mm Petri-dish
GS60-M
#1 1x3in. chamber slide and #1 60mm Petri-dish
6035-M
#1 60mm Petri-dish and #1 35mm Petri-dish
H201-MW-HOLDER
Multiwell plates holder
Figure 2.Available sample holders.
3. Available Lids
The following glass Lids are available for H201-NIKON-TI-S-ER:
•H201-SLIDING-LID: For easy sample loading and pipetting –Included in code H201-NIKON-TI-S-ER.
•H201-KOEHLER-LID*: Reduces chamber height to 21 mm and allows imaging under Koehler illumination.
•H201-INJECTION-LID*: Glass lid with two small openings (sealed with flexible plastic) allowing injection or
permanent access to the sample. Compatible only with sample holder 1x35.
•H201-LASER-INTERLOCK-LID*: Glass lid with safety switch. Connects to laser controller and automatically
turns laser off when lid is lifted. Compatible only with sample holder 1x35.
* OPTIONAL –not included with H201-NIKON-TI-S-ER

H201-NIKON-TI-S-ER User Manual
August 2019
www.oko-lab.com
Page5
3.1 Sliding lid
The Sliding Lid is a glass lid allowing for easy sample loading and pipetting. The Sliding Lid is screwed onto the cham-
ber.
Figure 3 shows chamber dimensions with the Sliding Lid (with and without chamber riser). Figure 4 illustrates how to
remove Sliding Lid, when a different Lid is necessary.
NOTE: Sliding Lid MUST BE REMOVED when using any other Lid. The Sliding Lid is fixed onto the chamber with 4
screws. Screws location is indicated by letter A in Figure 4 (image 2 and 3). Keep Sliding Lid fully closed to access
screws labeled A in image 2. Open Sliding Lid to access screws labeled A in image 3.
Sliding Lid
Distance between focal plane
and upper side of the glass
lid without the chamber riser
Distance between focal plane
and upper side of the glass
lid with the chamber riser
Chamber top view
Figure 3. H201-NIKON-TI-S-ER (Sliding Lid comes as standard)
1 2 3
Sliding Lid
A
A
Screws M2x4
Figure 4. Assembly of the sliding lid

H201-NIKON-TI-S-ER User Manual
August 2019
www.oko-lab.com
Page6
3.2 Koehler Lid
The Koehler Lid is a glass lid reducing the chamber’s height to 21 mm and allowing imaging under Koehler illumination.
The Koehler Lid is placed onto the chamber (NO SCREWS or TOOLS are necessary).
Figure 5 shows chamber dimensions with Koehler lid (with and without chamber riser).
Distance between focal plane
and upper side of the glass
lid without the chamber riser
Distance between focal plane
and upper side of the glass
lid with the chamber riser
Chamber top view
H201-KOHLER LID
Figure 5.H201-NIKON-TI-S-ER + H201-KOEHLER-LID
3.3 Injection Lid
The Injection Lid is a glass lid with two small openings sealed with flexible plastic allowing injection or permanent
access to the sample. NOTE: Compatible with sample holder 1x35 ONLY. The Injection Lid is placed onto the chamber
(NO SCREWS or TOOLS are necessary).
Figure 6 shows chamber dimensions with the Injection Lid (with and without chamber riser).

H201-NIKON-TI-S-ER User Manual
August 2019
www.oko-lab.com
Page7
Distance between focal plane
and upper side of the glass
lid without the chamber riser
Distance between focal plane
and upper side of the glass
lid with the chamber riser
Chamber top view
H201-INJECTION LID
Figure 6. H201-NIKON-TI-S-ER + H201-INJECTION-LID
3.4 Laser Interlock Lid
The Laser Interlock Lid is a glass lid with a safety switch connecting to the laser controller. It automatically turns the
laser off when the lid is lifted. NOTE: Compatible with sample holder 1x35 ONLY. The Laser Interlock Lid is placed
onto the chamber (NO SCREWS or TOOLS are necessary).
Figure 7 shows chamber dimensions with the Laser Interlock Lid (with and without chamber riser).
Distance between focal plane
and upper side of the glass
lid without the chamber riser
Distance between focal plane
and upper side of the glass
lid with the chamber riser
Chamber top view
H201-LASER-INTERLOCK LID
Figure 7. H201-NIKON-TI-S-ER + H201-LASER-INTERLOCK-LID

H201-NIKON-TI-S-ER User Manual
August 2019
www.oko-lab.com
Page8
4. Insertion of the Sample Feedback Temperature Sensor
Insert the Sample Feedback Temperature Sensor through the dedicated opening located in the H201-NIKON-TI-S-ER
(see Figure 8, Front and 3D views).
1.Frontal view 2.3D view
Temperature Sensor dedicated opening
Temperature Sensor dedicated opening
Figure 8. Insertion of the temperature sensor inside the chamber.
5. Insertion of Sample Holder into Chamber
Sample holders fit into the chamber base and are held in place by magnets embedded within both chamber and hold-
er. To introduce a sample holder with the proper orientation, match the red dot on the holder to the one on the
chamber base, as illustrated in Figure 9.
Insert the adapter by
superimposing the red points
Figure 9. Introduction of the Specimen Holder inside the Chamber Base.

H201-NIKON-TI-S-ER User Manual
August 2019
www.oko-lab.com
Page9
6. Working with 35 or 60 mm Petri Dish –Spacing Rings and
Magnetic Locks
Magnetic locks prevent movement of 35 and 60 mm dishes inside the sample holder.
Figure 10 illustrates the available magnetic locks for 35 and 60 mm dishes. Threaded magnetic posts allow adjusting
holder’s height.
Magnetic Locks for 35 and 60 mm petri dishes.
Included in the corresponding sample holder
Figure 10.Magnetic locks for 35 and 60 mm dish
NOTE: Magnetic locks are included with sample holder.
Spacing rings accounting for variability in dish diameter from one manufacturer to another are also included. Figure
11 shows how to position the ring. Ring selection guide diagrams: rings for 35 and 60 mm dish -Figure 12 and Figure
13, respectively.
1
3
2
Figure 11. 1) Insertion of the ring, 2) insertion of the 35 mm dish, 3) insertion of the magnetic lock.
NOTE: Spacing rings are included with sample holder.
NOTE: Spacing rings do not block the petri inside its accommodation, but they are designed to center the petri with
respect to the observation circular window.

H201-NIKON-TI-S-ER User Manual
August 2019
www.oko-lab.com
Page10
BD Falcon-35x10 Willco-35x10
Grainer-Petri35x10 Ibidi μ-Dish 35 mm-low
MatTek-P35G-X14-XCorning-35x10 mm
Figure 12. Ring selection scheme for 35 mm dish.
Corning-60x15 mm
BD Falcon-60x10
Ibidi μ-Dish 50 mm-low
Grainer - Petri 60x15
MatTek-P50G-X30-X
Willco 60x15
Figure 13. Ring selection scheme for 60 mm dish.

H201-NIKON-TI-S-ER User Manual
August 2019
www.oko-lab.com
Page11
7. Working with 1x3’’ and 1x2’’ chamber slides - magnetic
locks
Magnetic locks prevent movement of 1’’x 3’’ and 1’’x 2’’ chamber slides inside of the sample holder.
NOTE: Magnetic locks are included with sample holder.
1
2
Figure 14. Magnetic lock for 1'' x 3'' chamber slide.
1
2
3
Figure 15. Magnetic lock for 1'' x 2'' chamber slide. To lock the slide, push simultaneously the buttons indicated with (3).

H201-NIKON-TI-S-ER User Manual
August 2019
www.oko-lab.com
Page12
8. Working with MW Plates - Magnetic locks and Chamber ris-
er
Magnetic locks hold MW plates in place into sample holder illustrated in Figure 16
NOTE: MW plates holder must be ordered separately (NOT INCLUDED). Product code: H201-MW-HOLDER
Figure 16. Magnetic locks for MW plates. To be ordered separately.
Chamber Riser increases chamber’s height and is REQUIRED with 6, 12, 24, 48 MW plates, regardless of the Lid em-
ployed.
Figure 17 shows how to install and fasten the chamber riser in place. Screws location is indicated by letter A.
A
Chamber Riser
A
Chamber Base
Figure 17. Riser Assembly.

H201-NIKON-TI-S-ER User Manual
August 2019
www.oko-lab.com
Page13
9. Connection of the Gas Supply
A single silicon tubing carries output gas from the Okolab Gas Controller to the H201-NIKON-TI-S-ER. Silicon tubing
connects to a gas input - brass opening - located on a corner of the H201-NOKON-TI-S-ER. See Figure 18. Connect by
gently pushing silicon tubing onto brass opening.
Gas Input
Figure 18. Connection with gas supply.
10. Working with Perfusion
The Chamber Riser included with H201-NIKON-TI-S-ER features 12 perfusion holes for the insertion of perfusion tubing
up to 2.5 mm in outer diameter. To lock the Chamber Riser into the base use a 1.5mm Allen key to secure the 4
GUTB2-2,5-6 screws as shown in Figure 19.
Figure 19. How to lock the Chamber Riser.
Small screws plug the perfusion holes when not in use. (Grub screws M3x6). Remove small screws with a 1.5 mm Allen
key as necessary before introducing perfusion tubing.
Figure 20 shows location of perfusion holes.

H201-NIKON-TI-S-ER User Manual
August 2019
www.oko-lab.com
Page14
1.Frontal view 2.3D view
Perfusion dedicated opening
Figure 20. Perfusion.
Figure 21. How to access the perfusion holes.

H201-NIKON-TI-S-ER User Manual
August 2019
www.oko-lab.com
Page15
MICROFLUIDIC-C-RISER (Optional)
The MICROFLUIDIC-C-RISER is a chamber riser with 19x3 mm windows (on the two short sides) to allow insertion of
microfluidic tubings. Windows are magnetically closed. Compatible with most of H201 and H301 Okolab chambers.
Thickness of this chamber riser is 8mm (2mm more than default chamber riser). Requires Koehler lid and single ac-
commodation specimen holders.
Figure 22. MICROFLUIDIC-C-RISER.

H201-NIKON-TI-S-ER User Manual
August 2019
www.oko-lab.com
Page16
11. Connection of the Chamber with adapter
Follow the steps shown in the images of Figure 23 and listed below in order to correctly install the chamber onto XY
stage:
1. Follow the arrows shown in Figure 23 Images 1 and 2 for placing the captive screws in the proper housings
2. Place the chamber on the adapter as shown in Figure 23 Image 3
3. Use a 1.5mm metric Allen Wrench to tighten two of the four captive screws in place (A in Image 4) while keeping
the sliding lid closed. (See Figure 23 Image 4)
4. Tighten the remaining two captive screws (A in Image 5) while keeping the sliding lid open.
5. Image 6 of Figure 23 shows the chamber fixed on the adapter
1 2
34
56
Captive Screw
A
A
Figure 23. How to assemble the chamber to the adapter.

H201-NIKON-TI-S-ER User Manual
August 2019
www.oko-lab.com
Page17
12. Connection of the Chamber with XY stage
Once you have assembled the chamber with adapter you can place it on the XY stage, following the steps
shown in Figure 24.
1. Place the adapter on the stage (See Figure 24 Image 1).
2. Use the Grub screws to align correctly the adapter on the stage. Grub screws housings are indicated with letter A
in Image 2 of Figure 24.
3. Follow the arrows shown in Image 3 of Figure 24 for placing flat socket head cap screws; use the provided Allen
wrench for tightening the flat socket head cap screws and fastening the adapter on the stage. Screws housings
are indicated with letter B in Image 4 of Figure 24.
1 2
A
A
B
B
4
3
Screw M4x7
Screw M3x3
Figure 24. How to fix the H201-NIKON-TI-S-ER to the XY stage.