Laser Scanning Imaging System

What is the Laser Scanning Imaging System?

A laser scanning imaging system moves a focused beam point by point across a blot, gel or storage phosphor screen and records the signal with a photomultiplier, giving high spatial resolution and a very wide linear dynamic range for quantitative densitometry.

Overview

The scanner raster-scans a laser across the sample and detects emitted or transmitted light with a photomultiplier tube, building the image pixel by pixel. Because excitation is tightly focused and detection is fast, even weak signals are recorded with little noise, and the readout is linear over four to five orders of magnitude, which suits purity determination where small impurity peaks must be measured accurately. Formats include fluorescence scanning with multiple excitation wavelengths, storage phosphor imaging for radioisotopes, and densitometry of stained gels with transmitted light. Resolution can be set from 10 to 200 µm, so large blots can be scanned quickly at low resolution and re-scanned at high resolution for detailed work. Scan times range from one to thirty minutes depending on resolution and area.

How does it work?

Imaging systems convert a signal on a membrane or gel surface into a calibrated digital image. In chemiluminescence imaging, no excitation light is needed: an enzyme such as horseradish peroxidase converts a substrate into an excited product that emits light, and a cooled CCD or CMOS sensor integrates that faint emission over minutes. Fluorescence imaging instead excites fluorophores with LEDs or lasers and collects emitted light through bandpass filters, which allows multiplexing with differently labelled antibodies. Colorimetric systems use white or coloured illumination and measure transmitted or reflected light to quantify stained bands such as Coomassie blue. Laser scanning instruments move a focused beam across the surface and record point by point, which gives high spatial resolution and a wide linear dynamic range. In all cases the software subtracts background, converts pixel intensities to optical density, and compares bands with a standard or loading control.

What are the key features?

• Point-by-point scanning with photomultiplier detection
• Linear dynamic range of four to five orders of magnitude
• Selectable resolution from about 10 to 200 µm
• Multiple fluorescence excitation wavelengths
• Storage phosphor mode for isotopic detection
• Transmitted light densitometry for stained gels

What are the technical specifications?

A typical laser scanning imaging system configuration offers resolution of 10–200 µm selectable, dynamic range of approx. 4–5 orders of magnitude, excitation wavelengths of 488, 532, 635 and 685 nm options, detection of Photomultiplier tube with emission filters, scan area of approx. 250 x 250 mm and scan time of approx. 1–30 min depending on resolution. Values are indicative of the product class; the exact figures for each configuration are confirmed on the quotation and the calibration documents supplied with the instrument.

Which applications is it used for?

The Laser Scanning Imaging System is used in accurate purity determination of stained gels, radioisotope-labelled blot and gel imaging, multiplexed fluorescence quantification and high-resolution documentation of large blots. Across the wider Protein Imaging class, laboratories also deploy it for western blot band quantification, protein purity assessment from stained gels and multiplexed fluorescence immunodetection.

Which standards and certifications apply?

Relevant standards and methods include USP <1057>, Ph. Eur. 2.2.31, IEC 61010-1, ISO 17025, 21 CFR Part 11 and ISO 9001. Certification candidates for this model are CE, ISO 9001 manufacturing and IEC 61010-1 safety; each is to be confirmed for the specific configuration before shipment, and certificates are supplied only where they have been issued.

Keep the imaging area clean and free of dust, because particles scatter light and appear as false bands. Avoid saturating the sensor, since saturated pixels cannot be used for quantification; most software flags them, and exposure series should be used to confirm the signal is in the linear range. Chemiluminescent substrates decay within minutes to an hour, so imaging should follow the supplier's timing recommendation and be consistent between samples. Fluorescent blots fade with repeated exposure, so images should be captured once where possible. Calibrate the instrument periodically with a step tablet or fluorescence standard and record the settings used, so that images from different sessions remain comparable. Archive raw files, not only processed figures, because quantitative results may need re-analysis.

How do I select the right Laser Scanning Imaging System?

Select a laser scanning system when a very wide linear range, high resolution or isotopic detection is required, such as purity testing with small impurity peaks. Specify excitation wavelengths, resolution range, scan area, mode options and the quantification software included.

How does it compare with other Protein Imaging variants?

Choose the detector that matches your detection chemistry. Chemiluminescence needs a cooled sensor with low dark current and long integration times, and absolute sensitivity matters more than resolution. Fluorescence needs matched excitation sources and emission filters, and filter selection should cover the dyes you actually use, including near-infrared pairs for low-background multiplexing. Colorimetric and densitometry work needs uniform illumination and colour calibration to make Coomassie or silver staining quantitative. Laser scanning suits applications where a wide linear dynamic range and high resolution are essential, such as purity determination with small impurity peaks. Check the camera or scanner resolution in millimetres per pixel, dynamic range in orders of magnitude, cooling temperature, imaging area, light source lifetime and whether the software supports background subtraction, saturation warnings, lane profiling and export of numerical data.