Overview
The pE-4000 sets the standard as the universal Illumination System for fluorescence microscopy. The system has 16 selectable LED wavelengths across four channels that can be finely controlled and matched to the filters and fluorophores of almost any microscope, making it the broadest and most versatile illumination system available.
The CoolLED pE-4000 benefits from our award winning sustainable Green technology and delivers enhanced intensity at the sample plane with a significant reduction in energy consumption, and is supplied with a 36 month warranty.
The pE-4000 is a powerful, flexible, controllable system for advanced research applications.
“Striking, bright fluorescence images with a strong signal to noise ratio even when using a low magnification objective”
Graham Wright, Institute of Medical Biology, A*STAR Singapore
Advanced control for cutting-edge research
Multiple levels of control are possible, from simple to advanced. A dual mode control pod gives immediate manual control in either White or Advanced modes, where individual LED channels can be individually switched on/off or intensity modulated.
Alternatively, the USB interface and integration into major microscope software platforms allows complex software control.
TTL control enables capturing high-speed events (<1 ms). Since there is no shutter or filter wheel, without these moving parts, this reduced latency means speed is limited only by the camera. An expansion box is also available for added functionality with multiple BNC connections for remote analogue and TTL control.
For optogenetics applications, alongside the ability to include excitation filters to tune the wavelength to the opsin’s maximum activation peak, an internal function generator controls pulse timing and waveforms.
Improving cell viability
With the pE-4000, it is now possible to prevent artefacts and allow live samples to thrive in experiments >100 hours. Fine control of illumination timing and intensity with TTL and the function generator minimises exposure, avoiding photobleaching and phototoxicity.
“When you can only control the intensity of ‘white’ light (rather than individual channels), the level of photobleaching can be high. With the pE-4000, we can control the excitation of the individual channels. It is possible to optimise the excitation intensity according to the labelling, greatly reducing photobleaching and phototoxicity in a live experiment.”
Dr Yan Gu, University of Sussex
“Suddenly, we were able to offer users uninterrupted extended live cell experiments of 100+ hours, without worrying about brightness fluctuations, lamps burning, room heating, etc. Also, users have reported markedly reduced bleaching and phototoxicity in their samples, both from the prokaryotic and the eukaryotic research fields.”
Dr Jens Eriksson, Oslo University Hospital
Matching diverse fluorophore requirements
Full filter compatibility and 256 wavelength combinations from 16 installed LEDs (365 nm -770 nm) covers all available fluorophores, which is ideal for multi-user facilities or evolving experimental needs.
Multi-channel experiments with no pixel shift
Up to four channels using distinct wavelengths can be captured with no mechanical movement, preventing pixel shift and allowing the direct comparison of data as in the following experiment:
(A) Representative photocurrent traces of a PhobosCA expressing CA1 cell evoked with different activation wavelengths and shutoff with 595 nm light. (B) Photocurrent traces in the same cell evoked with 460 nm light and shutoff with indicated wavelengths (10 mW/mm2).1
Increased power efficiency
Our unique Wavelength Grouping Concept makes it possible to deliver more power in an efficient four-channel system. As the first company to introduce LEDs into fluorescence microscopy, CoolLED has developed a comprehensive understanding of the complexities of configuring and selecting filter sets for experiments using multiple fluorophores. CoolLED’s innovation comes from recognising that all the fluorophores used in multi-band work can have their absorption bands divided into four separate groups across the spectrum due to the restricted bandwidth available.
DAPI/FITC/TRITC/Cy5 Quad filter excited by pE-4000 at matched wavelengths shown above.
Performance
- Broadest LED spectrum available: 365-770 nm, with 16 LEDs
- Four channel wavelength-grouping design matches most commercially available dual, triple, and quad filters
- Removable inline excitation filter holders: no moving parts for fast acquisition
- Internal function generator for electrophysiology and optogenetics applications
- TTL and USB interfaces with major imaging software packages allows incorporation into all experimental parameters
- Analogue inputs for dynamic intensity control
- Individual control of selected LED wavelengths reduces background
- Rapid switching between LED wavelengths enables capture of high-speed events
- Instant on/off: No shutters required, no warm up or cool down
- Stable and repeatable: reliable and consistent results
- Precise intensity control in 1% steps (0-100%): no ND filters required
Convenience
- Pre-sets allow lab manager to match white spectrum to existing filter cubes
- Simple to fit, simple to use: no alignment, a once only adjustment
- Wide range of microscope adaptors: fits most microscopes
Plus all the benefits of LED technology.
The system comprises a pE-4000 Light Source, Control Pod, liquid light guide, Universal Collimator and Power Supply.
Control
Manual:
Dual function remote manual control pod for White mode or Advanced mode
Remote:
Via USB for independent on/off and intensity control of each channel. Triggering speed approximately 10-20 ms
Via 4 TTL inputs for independent on/off control of each channel. Triggering speed <20 µs
Via single TTL for on/off control of manual or software selected channels
Via 4 analogue inputs 0-10 V, 0-300 kHz for dynamic control of intensity from external analogue signals
Sync Out:
4 TTL outputs for each channel – active high 1 TTL output for any channel – active high
Programmable interface:
4 TTL outputs for on/off control of peripherals (transmitted illuminators, stages etc.) 4 analogue outputs for intensity control of peripherals (can be programmed to mirror LED intensities for channel control) 0-10 V full scale.
Function Generator:
Internally generated sine, pulse and ramps for each channel programmed via control pod.
Connectivity:
USB (B type) for PC connection. All other TTL and Analogue inputs/outputs via 25-way ‘D-type’ female connector (optional rear mounting pE-Expansion Box available for BNC connectivity).
Imaging Software:
Recognised under common software e.g. Micromanager, MetaMorph, cellSens, NIS Elements, etc. see Imaging Software
Choose liquid light guide or fibre
The pE-4000 can be configured to deliver light to the microscope via liquid light guide or fibre. Liquid light guides can be used in conjunction with CoolLED’s pE-Universal Collimator which can accept a microscope adaptor from CoolLED’s extensive range. Find out more here.
Specification
Power requirements
110-240 V a.c. 50/60 Hz, 2.5 A
Power consumption
Standby (i.e. no LEDs on) : Max 7 W
Single wavelength operation : Max 25 W
Dual wavelength operation : Max 44 W
Triple wavelength operation : Max 53 W
Quad wavelength operation : Max 60 W
Dimensions
pE-4000 Light Source: 150 mm(w) x 22 0mm(d) x 260 mm(h) – Weight 3.5 kg
pE-4000 Control pod: 154 mm(w) x 135 mm(d) x 40 mm(h) – Weight 0.95 kg
pE-4000 Power Supply: 164 mm(w) x 64 mm(d) x 35 mm(h) – Weight 0.58 kg
pE-Expansion Box: 151 mm(w) x 18 mm(d) x 95 mm(h) – Weight 0.34 kg
Environment and Safety
- Mercury-free and Laser-free
- Energy Efficient
- Long lifetime
- No bulb replacements
- Reduced risk of eye damage
- Quiet operation
- No special disposal regulations or issues
Warranty System is 36 months
I have been using a pE-4000 for optogenetic experiments using in vitro slices for just over a year now. The pE-4000 has been fantastic for my needs: the ability to simultaneously use multiple wavelengths of light has allowed me to carry out experiments using two different opsins to study how external inputs converge on hippocampal interneurons. I’m currently setting up my own research group at the University of Exeter, and all of my patch-clamp rigs will have a pE-4000 on them, as they are such versatile devices for optogenetic experiments.
Dr Michael Craig, Research Fellow, University of Exeter Medical School
Our group is currently attempting to isolate and stain circulating tumour cells. In order to identify these cells we need a minimum of four colour fluorescence imaging, requiring the slide to be scanned using four different LEDs.To stitch our images together after scanning we require a transmission light microscope image. The image co-ordinates of the light microscope image are used to stitch the fluorescence images together. The previous light source (PE-2) allowed only three LEDs to be used together for our purposes. This meant that when we have multiple slides there are two options to get the full four fluorescence scan:1) Change the LED after scanning in Brightfield and three fluorescence channels – this risks damage to the LED itself and is time consuming. It involves un-screwing a panel each time and screws can be damaged or lost relatively easily. It places demand on the ribbon that is connected to the LED and, over time, may damage the ribbon also. We usually scan multiple slides and prefer to use option two, to reduce wear and tear on the light source.2) This option is to scan all slides in Brightfield and three colour fluorescence and then go for a second batch of images in both Brightfield and the final fluorescence channel. This is also time consuming, as setting up each scan in focus takes time. It also requires a lot more downstream processing to line the images up with one another, as the scans are often of two different sizes.With the new PE-4000 light source I was able to scan four slides in Brightfield and four fluorescence channels in three hours. Four slides took six hours to scan on the previous light source, due to double the number of scans and changing the LED. It has also reduced the downstream processing of the images, as they are all taken using the same co-ordinates, in one batch.Our project aims to expand from four colour fluorescence to six or seven channels over the coming years. Using the old light sources this would require a minimum of three but possibly four scans taken from different positions per slide and would take hours per slide.The current light source meets the needs of researches using modern immunofluorescence techniques as most projects have moved from single to a multi-marker approach, in recent years.
Anthony Cooney, PhD Student, Department of Histopathology, Trinity College Dublin.
I have been imaging using the new CoolLED PE-4000 system on our epi-fluorescence microscope. This is an excellent system as it covers a very wide spectrum of wavelengths. I have found it very easy to use. The user interface is much better as the different LEDs can be changed using the handheld control- without switching off and unscrewing the LAMs as had to be done on the old PE-2 system. In addition the LEDs can be changed dynamically while the software to capture the image is open, and also a huge combination of different wavelengths can be used at the same time.I have found that the light source from this system is considerably stronger than from the previous system - I now image at 50% strength, when I used to image at 100%.I anticipate using this imaging system a lot more now that the new LED light source is there.
Alison Reynolds, PhD Research Fellow, UCD Conway Institute, University College Dublin
I am very impressed with the brightness and performance of the CoolLED pE-4000 in comparison to the mercury or halide light sources for fluorescence. I also really like its neat packaging and it is easy to set up and configure. Absolutely brilliant upgrade for archaic mercury set ups!
Dominic White, Area Sales Manager, Carl Zeiss Microscopy GmbH
Throwing out the old mercury lamp and exchanging it for the pE-4000 on one of our live cell wide field microscopes has been an astounding success! It allowed us to cheaply breathe new life into old equipment. Suddenly, we were able to offer users uninterrupted extended live cell experiments of 100+ hours, without worrying about brightness fluctuations, lamps burning, room heating, etc. Also, users have reported markedly reduced bleaching and phototoxicity in their samples, both from the prokaryotic and the eukaryotic research fields. As a facility manager I am pleased with the instrument's ease of use, and shallow learning curve for new users. Taken together with reduced overall microscope maintenance, the pE-4000 has already saved me a substantial amount of time in my daily work life, and improved the quality of our services at the same time.
Dr. Jens Eriksson, Manager, Superresolution Microscopy Core Facility, Dep. Clinical Biochemistry, Oslo University Hospital.
Conceptually, the pE-4000 seems an extremely versatile tool that blends seamlessly with our range of microscopes, making it ideal for high end life science applications. It is also very user friendly and powerful.
Dr Alex Gardiol, PhD, Imaging Specialist, Olympus Microscopy
I just purchased a pE-4000 to replace an old version of Lumencor Sola SE on a live imaging microscope. After comparing the power level of the systems, I am convinced that the pE-4000 provides stronger illumination at the objective. As the Sola light source can only control intensity of ‘white’ light (rather than individual channels), we have to choose the excitation level according to the dimmest labelling fluorophore. The other channels are in turn exposed to an unnecessary high dose of excitation. The level of photo-bleaching can be high as a result. With the pE-4000, we can control the excitation of the individual channel. It is possible to optimise the excitation intensity according to the labelling, greatly reducing photo-bleaching and photo-toxicity in a live experiment. Due to the difference in the excitation spectrum with the Sola, I was worried about the extra cost of changing the dichroic, excitation and emission filters. The options of four wavelength in each channel (totally four channels) enables me to pick up the correct wavelength according to the old filter/dichroic settings.
Dr. Yan Gu, Manager, Analytical and Quantitative Light Microscopy Facility, Genome Damage and Stability Centre, University of Sussex
- Self-organized centripetal movement of corneal epithelium in the absence of external cues
- Appetite controlled by a cholecystokinin nucleus of the solitary tract to hypothalamus neurocircuit
- The C9orf72 protein interacts with Rab1a and the ULK1 complex to regulate initiation of autophagy
- Anion-conducting channelrhodopsins with tuned spectra and modified kinetics engineered for optogenetic manipulation of behaviour
- Choice of Illumination System & Fluorophore for Multiplex Immunofluorescence on FFPE Tissue Sections
A keratinocyte undergoing mitosis in which the keratin is labelled with GFP. Acquired on a Olympus IX-83 microscope – CoolLED pE-4000. Scale bar = 10 μm.
CoolLED pE-4000 working with cultured hippocampal neurons
Fluorescence image of 15 zebrafish larvae (fixed and stained with acridine orange) within the field of the Mesolens.
Time series of a confluent sheet of keratinocytes migrating during a live-cell imaging outgrowth assay, acquired on an Olympus IX-83 microscope equipped with a CoolLED pE-4000 lightsource (Fig. 4). Scale bar = 150 μm.
Structured illumination superresolution microscopy (3D-SIM) images of the GFP labelled keratin within (A) a wildtype and (B) a mutant keratinocyte cell. Acquired on a GE Healthcare DeltaVision OMX Blaze microscope. Scale bar = 10 μm.
Improving laboratory sustainability
Configuring Olympus cellSens
CoolLED Optimised filters
Cell Viability, Image Quality & Sustainability
Using Optogenetics to Analyse Neuronal Circuits
| Light Source | Emission Colour | Type | CoolLED LED | Chroma | Semrock | |||
|---|---|---|---|---|---|---|---|---|
| pE-4000 | Blue | Single | 365 | 49053 CoolLED DAPI | 49000 ET Dapi | LED-DAPI-B | ||
| pE-4000 | Blue | Single | 385 | 49053 CoolLED DAPI | 49028 ET Dapi 395 | LED-DAPI-B | CFW-BP01- Clinical | |
| pE-4000 | Blue | Single | 405 | 49053 CoolLED DAPI | 49028 ET Dapi 395 | LED-DAPI-B | ||
| pE-4000 | Cyan | Single | 435 | 49001 ET eCFP | 49013 ET Teal | LED-CFP-A | LED-mTFP-A | |
| pE-4000 | Green | Single | 460 | 49054 CoolLED GFP | GFP-4050B | GFP-A-Basic | ||
| pE-4000 | Green | Single | 460 | 49002 ET eGFP | 39002 AT eGFP | GFP-4050B | GFP-A-Basic | |
| pE-4000 | Yellow-Green | Single | 490 | 49303 ET Green#1 | 49003 ET eYFP | YFP-A-BASIC | ||
| pE-4000 | Orange | Single | 550 | 49004 ET Cy3 | 49005 ET dsRed | TxRed-4040C | LED-TRITC-A | |
| pE-4000 | Red | Single | 580 | 49055 CoolLED mCherry #1 | 49056 CoolLED mCherry #2 | TxRed-4040C | LED-mCherry-A | |
| pE-4000 | Red | Single | 580 | 49008 ET mCherry | 39010 AT Tex. Red | YFP-2427B | LED-mCherry-A | |
| pE-4000 | Red | Single | 595 | E9015 ET Alexa 633 | SpRed-B | |||
| pE-4000 | Far Red | Single | 635 | 49006 ET CY5 | 39007 AT Cy5 | Cy5-4040C | ||
| pE-4000 | IR (660) | Single | 660 | 49022 ET Cy5.5 | Cy5.5-C | |||
| pE-4000 | IR (740) | Single | 740 | 49007 ET Cy7 | 49037 ET LICor 800 | IRDYE800-33LP-A | Cy7-B | |
| pE-4000 | IR (770) | Single | 770 | 49030 ET Indocy. Gr. | ICG-B | |||
| pE-4000 | Blue-Green-Red-Far Red | Multiband | 405, 490, 550, 635 | (365, 385, 405), 460, 580, 635 | 89402 ET DAPI/FITC/TRITC/Cy5 | LED-DA/FI/TR/Cy5-B | ||
| pE-4000 | Blue-Green-Red | Multiband | (365,385,405), 460, 550 | (365, 385, 405), 460, 580 | 69401 – ET Triple Multi LED | LED-DA/FI/TX-A | ||
| pE-4000 | Blue-Green-Red | Multiband | 385, 490, 550 | 385, 460, 550 | 69000 ET DAPI/FITC/TRITC | DA/FI/TR-A | ||
| pE-4000 | Blue-Green-Red | Multiband | 385, 490, 580 | 69015 ET DAPI/Green/RedFISH | ||||
| pE-4000 | Blue-Green-Red | Multiband | 405, 490, 550 | 69010 ET DAPI/FITC/Cy3 | ||||
| pE-4000 | Blue-Green-Red | Multiband | 405, 490, 550 | 405, 490, 580 | 69002 ET DAPI/FITC/Texas Red | DA/FI/TX-B | ||
| pE-4000 | Cyan, Green, Orange | Multiband | 435, 490, 550 | 69011 ET Aqua/Green/Orange FISH | ||||
| pE-4000 | Cyan, Green, Red | Multiband | 435, 490, 580 | 435, 490, 580 | 69008 ET eCFP/eYFP/mCherry | LED-CFP/YFP/mCherry-A | ||
| pE-4000 | Blue-Green | Multiband | 405, 490 | 385, (460, 490) | 59001v2 ET DAPI/Green FISH | DA/FI-A | ||
| pE-4000 | Blue-Orange | Multiband | 405, 550 | 59002v2 ET DAPI/Orange | ||||
| pE-4000 | Blue-Red | Multiband | 405, 580 | (460, 490), 580 | 59003 v2 ET DAPI/Red FISH | FITC/TxRed-A | ||
| pE-4000 | Cyan-Yellow | Multiband | 435, 490 | 405, 490 | 59017 ET eCFP/eYFP | CFP/YFP-A | ||
| pE-4000 | Aqua-Gold | Multiband | 435, 550 | 59033 ET Aqua/Gold FISH | ||||
| pE-4000 | Green-Red | Multiband | 460, 580 | 460, 550 | 59022 ET eGFP/mCherry | GFP/DsRed-A | ||
| pE-4000 | Green-Red | Multiband | 490, 580 | 59009 ET FITC/Cy3 | ||||
| pE-4000 | Green-Red | Multiband | 490, 580 | 59010 ET Green/Red FISH #1 | ||||
| pE-4000 | Blue-Green-Red-Far Red | Pinkel | 365, 405, 490, 550, 635 | (365, 385, 405), 460, 580, 635 | 89400 ET DAPI/FITC/TRITC/Cy5 | LED-DA/FI/TR/Cy5-4X-B | ||
| pE-4000 | Blue-Green-Red | Pinkel | (385, 405), 460, 550 | (365, 385, 405), 460, 580 | 89084v2 ET DAPI/Green/Orange FISH | LED-DA/FI/TX-3X-B | ||
| pE-4000 | Blue-Green-Red | Pinkel | 405, 490, 550 | 385, 460, 550 | 69300 ET DAPI/FITC/TRITC | DA/FI/TR-3X-A | ||
| pE-4000 | Blue-Green-Red | Pinkel | 385, 490, 580 | 89085 ET DAPI/Green/Red FISH | ||||
| pE-4000 | Blue-Green-Red | Pinkel | 405, 490, 580 | 69302 ET DAPI/FITC/Texas Red | ||||
| pE-4000 | Cyan, Green, Red | Pinkel | 435, 490, 580 | 69308 ET eCFP/eYFP/mCherry | LED-CFP/YFP/mCherry-3X-A | |||
| pE-4000 | Cyan-Yellow | Pinkel | 435, 490 | 59217 ET eCFP/eYFP | CFP/YFP-2X-A | |||
| pE-4000 | Green-Red | Pinkel | 460, 580 | 490, 580 | 59222 ET eGFP/mCherry | FITC/TxRed-2X-B | ||
| pE-4000 | Green-Red | Pinkel | 460, 550 | 59204 ET FITC/TRITC | GFP/DsRed-2X-A | |||
| pE-4000-L-SYS-ZZ: | pE-4000 Light Source with manual control pod, and power supply for LLG delivery |
| pE-4000-F-SYS-ZZ: | pE-4000 Light Source with manual control pod and power supply for fibre delivery |
| pE-4000-EB25D: | Rear mounting pE-Expansion Box for 25-way D-type to BNC connectivity |
| pE-4000-EFH-4: | Set of four Excitation Filter holders (25 mm dia.) |
| pE-6501: | USB-TTL Conversion Kit |
| pE-1906: | 1.5 m long, 3 mm diameter liquid light guide |
| pE-10400-YYY: | pE-Universal collimator for use with a single liquid light guide and customer specified adaptor. |
We supply a range of fibres, available via the Accessories tab
To specify microscope code (YYY), see Adaptors
To specify Power Cable (ZZ): 10 = Australia, 20 = Europe, 30 = UK, 40 = USA
pE-Expansion Box
Fibres
Liquid Light Guide
Interchangeable Light Delivery Output module for Fibre (SMA/FC fittings)
Interchangeable Light Delivery Output module for LLG (3 mm diameter)
Filter Holders
pE-6501
The CoolLED pE Driver is first required for software integration, and can be downloaded here.
CoolLED Proprietary ZEN Bridge software is available – Contact CoolLED for further information.