UV Microscope for Protein Crystal Inspection

Trying to tell apart protein crystals from non-protein crystals in crystallization drops has been a longstanding problem in protein crystallography. The ultimate differentiator of course is the X-ray diffractometer. However, this also is impractical in many cases - the crystals may be too numerous, they may be too small, etc.

Look at this example of crystalline objects that are only a few micro-meters long:

Birefringence gave uncertain results. At this point, one option is to plow ahead and optimize the condition, use the microcrystals for seeding, etc., but is it worthwhile when time and resources could be spent on cases with more certain prospects for success?

Other examples that are useful for crystallographers are the case of phase separation - is the protein inside or outside the "oily" drops?
When there is precipitate in the crystallization, is it protein?

UV imaging, it turns out, can address these questions in situ in a rapid, non-destructive way.

The above example under UV fluorescence imaging:

An example of phase separation of drops ripe with protein:

Protein microcrystals mixed with protein precipitate:

References:

1. Judge, RA, et al. [2005] "An ultraviolet fluorescence-based method for identifying and distinguishing protein crystals", Acta. Cryst. D61, 60-66.
2. Lunde, S, et al. [2005] "UV microscopy at 280 nm is effective in screening for the growth of protein microcrystals", J. Appl. Crystal. 38, 1031-1034.
3. Gill, HS [2010] "Evaluating the efficacy of tryptophan fluorescence and absorbance as a selection tool for identifying protein crystals", Acta. Cryst. F66, 364-372.
Near UV (300 - 400 nm) fluorescence in proteins originates predeominantly from Trp and Tyr residues. By choosing an excitation wavelength that induces Trp and, to some extent, Tyr fluorescence, we can cover almost all protein molecules.

When it is present, fluorescence from Trp dominates over fluorescence from Tyr. Trp may also quench Tyr fluorescence. Trp fluorescence itself may be quenched by the presence of nearby charged residues, bound ligands and cofactors, due to energy or electron transfer. The fluorescence quantum yield of Trp in proteins varies from 0 to 40%. Hen egg white lysozyme, with 3 Tyr residues and 6 Trp residues has a near-UV fluorescence quantum yield of 12%, owing mainly to two Trp residues.

Prolonged excitation of Trp residues can result in photochemical conversion of the amino acid and lead to a loss of near-UV fluorescence. [Repeated excitation at shorter wavelengths can cause denaturing and/or degradation of the protein.]

To avoid damage to the protein crystals, excitation and emission must be very efficient. We achieve this in the UVEX by careful design of the optics, choice of optical elements and a very sensitive imager.
In the event where UV fluorescence is quenched due to the binding of a small molecule to the protein, UV transmission is a more dependable means of identifying the protein crystal. In fact, when the apo-protein is fluorescent and the complex is not, comparison of UV fluorescence image with UV transmission image provides a powerful method for detecting crystals of the complex.

UV transmission imaging also is very suitable for crystals of nucleic acids and does not require staining the crystals with fluorescent dye for identification. [See the Images section.]

UV transmission requires UV-transparent optics, filters different than needed for UV fluorescence, and a camera sensitive down to 280 nm.

The optics and camera of the UVEX are suited for UV transmission imaging. The camera is cooled to minimize the contribution of hot pixels. The change-up of filters and light source in absorbance option are implemented seamlessly so that it's a snap to switch between UV fluorescence and absorbance modes.

UVEX-m & UVEX-M - manual models
UVEX-P - automated model for plate scanning
UVEX-PS - automated model with plate storage (capacity 42 - 504 SBS plates)
Add an A at the end for Absorbance option

Common to all models

Dimensions 12" x 12" x 18"

Weight 30 lb

Electrical 110 - 240 V; 50 - 60 Hz; 600 W max

Light sources All LED, pulsed on-demand, instant on, instant off

Optics Nominally 5x and 15x objectives standard, other magnifications optional

Optical Resolution as low as 0.5 mm/pixel; coverage area 3 mm

Detector Cooled monochrome camera with high UV sensitivity

The UVEX is designed and manufactured in the USA

- Japan
  
  BioSpeed Co., Ltd.
  4F Ohsawa Bldg, 3-6-9
  Bunkyo-ward, Tokyo 113-0033
  Ph: +81-3-5840-9491
  website
- India
  
  Biotech Desk Pvt. Ltd
  4F 5, Ballad Estate
  Tarnaka, Hyderabad 500 017
  Ph: +91-40-2701-7477
  website
- China
  
  LBD Lifesciences Ltd.
  Room 301, Bldg 2, No 88, Darwin Road
  Zhangjiang High-tech park, Pudong District, Shanghai, 201203 N
  ph: + 86-21-51871506 / 51973610
  website
Contact

JAN Scientific, Inc.
4726 Eleventh Ave NE, Suite 101, Seattle, WA 98105, USA

Telephone: +1 206.632.1814
FAX: +1 206.632.4044
E-mail: admin@janscientific.com

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UV Microscope for Protein Crystal Inspection

UVEX-m & UVEX-M - manual models
UVEX-P - automated model for plate scanning
UVEX-PS - automated model with plate storage (capacity 42 - 504 SBS plates)
Add an A at the end for Absorbance option

The UVEX is designed and manufactured in the USA

Japan

India

China

Autoscanning in action

Brightfield, UV Fluorescence and UV Absorbance Imaging

Contact

JAN Scientific, Inc.
4726 Eleventh Ave NE, Suite 101, Seattle, WA 98105, USA

Events

Privacy Policy

Your email and other contact information will not be shared with or sold to a third party

	Dimensions	12" x 12" x 18"
	Weight	30 lb
	Electrical	110 - 240 V; 50 - 60 Hz; 600 W max
	Light sources	All LED, pulsed on-demand, instant on, instant off
	Optics	Nominally 5x and 15x objectives standard, other magnifications optional
	Optical	Resolution as low as 0.5 mm/pixel; coverage area 3 mm
	Detector	Cooled monochrome camera with high UV sensitivity

UVEX-m & UVEX-M - manual models UVEX-P - automated model for plate scanning UVEX-PS - automated model with plate storage (capacity 42 - 504 SBS plates) Add an A at the end for Absorbance option

The UVEX is designed and manufactured in the USA

Japan

India

China

Autoscanning in action

Brightfield, UV Fluorescence and UV Absorbance Imaging

Contact

JAN Scientific, Inc. 4726 Eleventh Ave NE, Suite 101, Seattle, WA 98105, USA

Events

Privacy Policy

Your email and other contact information will not be shared with or sold to a third party

UVEX-m & UVEX-M - manual models
UVEX-P - automated model for plate scanning
UVEX-PS - automated model with plate storage (capacity 42 - 504 SBS plates)
Add an A at the end for Absorbance option

JAN Scientific, Inc.
4726 Eleventh Ave NE, Suite 101, Seattle, WA 98105, USA