Laser Tweezers and Laser Scissors

Brian Hull Physics 461 March 21st, 2004

Laser Tweezers and Scissors Beginnings

Laser scissors came first
Donald E. Rounds and Michael W. Berns wrote a paper in Scientific American, february 1970 titled Cell Surgery by Laser.

Laser tweezers were first demonstrated by Arthur Ashkin (mid 1980s)

They performed optical trapping inside a cell without causing any damage to the cell wall or membrane

Early Laser Tweezers Experiments (Ashkin)

A continuous-wave (cw), low-power (<1W) laser was used to optically trap individual bacteria and protozoa First used a blue-green argon ion laser and second an infrared ND:Yag laser The experiments showed that organelles could be grabbed and moved around using the lasers

Early Laser Tweezers Experiments

Steven Chu from Stanford University showed that lasers could also manipulate molecules They demonstrated this by placing polystyrene beads at the ends of DNA and pulling on the beads to stretch the DNA Block of Princeton and Sheetz of Duke have used laser scissors to study kinesin motors (the structures that drive a flagellum or sperm tail)

Early Laser Scissors Experiments

Berns and his colleagues focused on defining the laser parameters and determining which organelles could be manipulated (~.25 m) Later found that laser scissors could be used to study organelles of the nucleus of a cell, including chromosomes, and the mitotic spindle Also began to study structures such as mitochondria, microfilaments & microtubules

Physics of Laser Tweezers

A small relatively transparent (to a certain laser frequency) object refracts incident beams, bending the light
This refraction transfers momentum from the light to the target object The momentum causes a Lilliputian force (pN) that pulls the object toward the focal volume of the beam
By moving the focal volume of the beam one can cause the object to move as well

Physics of Laser Tweezers

Force Measurements:
Force is linearly dependent on the laser power
F = QP/c
Where Q is a trapping efficiency parameter dependent upon the optical properties of the trapped object

Laser Tweezers Diagram

Trapping Beam

Fgrad

Cell

More about Laser Scissors

Important to achieve precision and selectivity
Precision refers to targeting
Due to technology (high level of optical elements in microscopes), precision is relatively straightforward

Selectivity refers to alteration of the target leaving the surroundings unaffected

Laser is a Gaussian distribution of energy

Energy at the focal spot is characterized by a bellshape curve
Only the peak of the curve may have sufficient energy to alter a target so the effective spot can be much less than the diameter of the measured focal spot

More about Laser Scissors

Unknowns about interactions between laser light and biological targets
Makes selectivity very difficult to guarantee without empirically noting what works

Differences between Laser Tweezers and Laser Scissors

Obvious
Cutting versus managing and manipulating

Scissors are short pulsed and high irradiance (energy reaching the surface in a given time) beams while tweezers are continuous and of low-irradiance Target MUST be transparent to tweezers in order not to pass significant energy

Side effects and Difficulties of Laser Tweezers and Scissors

Defining exact laser effects is difficult
Submicron interractions Energy controls in small volumes

Heat is a significant issue with Tweezers

The cw lasers can cause a 1.15 to 1.45 degree Celsius rise per 100 milliwatts of laser power If the dissipation is inefficient the laser can produce ten times that power

More Side effects and Difficulties

Can cause microplasma to form
A cloud of ionized gas that stresses the cell and can cause it to rupture Occurs when the laser is focused on or above the glass slide (of a microscope)

Single photon absorption may heat a target and can initiate chemical reactions resulting in free-radicals Photoablation
High energy photons (ultraviolet) can break molecular bonds, tearing molecules apart

Applications
Chromosome alteration
Scissors produce changes in chromosomes and can inactivate a selected chromosome Can cut and paste chromosomes during meiosis Leaves the chromosomes apparently unaffected Study mitotic spindle
Discovered that movement of fragments outside the spindle was easy but fragments in the spindle could not be moved

Applications cont
Optoporation
Cutting of micron sized holes in a cell
The cell closes the hole in a fraction of a second with no apparent permanent damage Within that time, molecules can be inserted into a cell

Suitable for genetic manipulation of plants especially (cell walls make manipulation difficult)

Exploit microplasma formation

A glass capillary tube collects the cell contents Applicable in single-cell analytical chemistry

Applications cont
UC Irvine: study immune system T and Bcells
T cells have a polarized appearance, a kind of front and back When B-cells were trapped and placed at the back of the T-cells, nothing occurred but when placed at the front, the immune system cascade began
(increasing Ca+ ions in T-cells and promoting their proliferation)

Applications cont
UC Irvine: Tweezers can grasp highly motile cells including male sperm cells
Study the swimming force of sperm Trapped sperm cells and analyzed their escape abilities
Discover zigzag pattern swimmers create a greater force than the straight pattern swimmers

Note however that exposure to tweezers can hamper motility of object or cell

Applications cont
Merging of two distinct cells to create a third cell with the genetics of both the previous two Manipulation of gametes
assisted hatching Scissors thin or remove a small area of the protective cover of an egg cell Fertilization by trapping and moving sperm cells to a pore in an egg created by laser scissors

Cutting Edge technology (pardon the pun)

National Institues of Health Laser Microbeam and Medical Program Biotechnology Resource
Built a microscope workstation with two laser tweezers and one laser scissors beam into a confocal laser fluorescence microscope Lasers are tunable to any wavelength
Researchers can observe fluorescing cells during and after use of laser tweezers and laser scissors Useful in studying DNA sequencing

References
Liang, Hong et al. Directed Movement of Chromosome Arms and Fragments in Mitotic Newt Lung Cells Using Optical Scissors and Optical Tweezers. Beckman Laser Institute and Medical Clinic, UC Irvine Berns, Michael. Laser Scissors and Tweezers. Scientific American. Apr98 V. 278, Issue 4 Konig, Karsten. Laser tweezers and multiphoton microscopes in life sciences. Robert Feulgen Prize Lecture. Springer-Berlag 2000. pub. On-line 19 July 2000 Fuhr, Gunter R. Living cells in opto-electrical cages. Department of Biology, Humboldt-University of Berlin, Invalidenstr. 42. Elsevier Science B.V. 2000 www.intracel.co.uk/cellrmws.htm

Questions?