Team:UANL Mty-Mexico/Contributions/Light Machine

Photobiological processes have been relevant in nature because they are found in plants, algae, cyanobacteria among other organisms that use light as an energy source, protection against radiation damage, regulatory and signaling systems, these organisms possess single/multiple photoreceptors, allowing them to adapt to their environment depending on the light. ^{1,4, 5}

Light is absorbed by specific chromophores, which are tuned, by their proteinaceous and external environment, to function optimally. All chromophores belong to three classes: tetrapyrroles, polyenes and aromatics. The chemical structure of photosensing pigment/protein complexes has been resolved for many of photobiological processes that have characteristic sensitivity in the visible and infrared part of the spectrum.^5,7

Under the tetrapyrroles class, are located the Phytochromes and Cyanobacteriochromes (bacteria-specific relative of the Phytochromes), which are used by Escherichia coli JT2 strain, red and green light-sensitive transcription system based on a red/far-red chimeric cyanobacterial phytochrome Cph1 and the E. coli EnvZ/OmpR component signaling pathway, plus the green/red CcaS cyanobacteriochrome and its response regulator CcaR.¹

Phytochromes are photoreceptors that typically perceive a specific wavelength in the visible/Infrared spectrum, regulate a wide range of physiological responses to light quality, quantity, duration, and direction. Cph8 and CcaS/CcaR exhibit reversible photoconversion two spectrally distinct forms, an active absorbing form and an inactive absorbing form respectively in their specific wavelengths. ^5,6

These two Phytochromes use the same phycocyanobilin chromophore (PCB) that binds at a conserved cysteine within an N-terminal GAF (cyclic GMP phosphodiesterase, adenylyl cyclase, FhlA) domain and imparts reversible photoactivation of signaling activity with maximal responses to 535 nm for green and 672 nm for red photoreceptor respectively.¹

This section explains extensively how to build a Light Machine!

Overview of Light Machine

We built a special machine that allows us to properly excite both photoreceptors (CcaS/CcaR and Cph8) independently/simultaneously using specific wavelength of 535 and 650 nm.

This "Light Machine" was based mainly on previous publications that work with light ^1,2,3

Materials

• Two 100 W halogen lamp (Philips)
• Green and Red bandpass interference filters
**We get ours from Edmunds Optics. Description: Interference Bandpass Filter 535nm 25mm Catalog Number: NT65-700.
http://www.edmundoptics.com/products/displayproduct.cfm?productid=3196&showall#products
And Interference Bandpass Filter 650nm 25mm Catalog Number: NT65-715
http://www.edmundoptics.com/products/displayproduct.cfm?productid=3197&showall#products
• 37°C incubator with thermometer hole on top
• Adjustable shelves
• Two Support Stands/Clamps
• Two Emisor-Receptor Pieces
• T-45° Block
• Aluminum tubes (33 mm*180 mm & 33 mm*90 mm)< mirror like finish inside
• Two mirrors 50 mm*50 mm
• Two porcelain sockets T4 base
• 2 Small Fans

Distance overview of "Light Machine"

Instructions

• First of all, collect or make all the materials listed above.
• Wallpaper completely the inside of the incubator with black paper.
• Clear the thermometer hole in the top of incubator such that light can pass through.
• Place the T-45° Block at the top of the incubator on the thermometer hole and fix it with a plastic ring. Place two mirrors on T-45°Block.
• Immobilize the bandpass filter inside a plastic 1' coupling with small plastic ring. Join the desire coupling to the small aluminum pipe of T-45° Block.
• Immobilize the emission-Receptor pieces at both sides of incubator, we use adjustable shelves.
• Join aluminum tube to Emision-Receptor.
• Leave proper distance * between coupling and aluminum pipe.
• Immobilize aluminum tubes with support stands.
• Place the aluminum tube in Emissor-Receptor piece and place the pipes parallel to the T-45° Block.

Turn on the lamps; adjust the positioning of mirrors such that a clear and homogeneous light pattern appears in the center of your incubator.

*Notes

****Have fans near Aluminum Pieces they come very hot over time, prevents halogen lamp and filter fails. Our incubator has a 1 3/4' hole When placing T-45° Block ensure to cover all light penetrable areas.
Buy more than 2 halogen lamps, they broke easily Be careful handling the halogen lamps, they heat in a very short time.
*Two finger rule

We buy 4 BP filters 532, 575, 650, 710 nm respectively Make experiments in dark room. Clean mirrors when possible. Measure light intensity before starting test.

Before starting making measurements it is necessary to understand the following concepts:

Radiometry: is the measurement of optical radiation from a physical point of view, includes the regions commonly called the ultraviolet, the visible and the infrared. Two out of many typical units encountered are Watt and Joule.

Photometry: is the measurement of visible light, which is detectable by the human eye. These measurements tend to be subjective. Typical photometric units include lumens, lux and candelas.

Photometry is almost the same as radiometry, except that radiometry includes the entire optical radiation spectrum, while photometry is limited to the visible spectrum as defined by the response of the eye.^8,10

Irradiance (a.k.a. flux density) is a SI derived unit and is measured in W/m2. Irradiance is power per unit area incident from all directions in a hemisphere onto a surface that coincides with the base of that hemisphere.

Illuminance (a.k.a. luminous flux density) is another SI unit and is measure in lux. Illuminance is the total luminous flux incident on a surface per unit area. (Is the photometric equivalent of irradiance).⁹

**In few words you need to measure in photometric units (lux) and convert them to radiometric units (W/m2) or measure directly in W/m2

To determine the light intensities once having assembled the "Light Machine", there are two types of devices to perform such tasks:

Photometer - is an instrument for measuring Illuminance (Photometric units), then this value will be converted into Irradiance (W/m2).

**Cheaper measurement device and works properly.

Spectrometer – is an instrument for measuring Irradiance.

** Is the device that has everything but it costs much more