Technical Features of the LightCycler® 2.0 Carousel-Based System
The innovatively designed LightCycler® 2.0 Instrument produces results in a fraction of the time required by other PCR methods.
They use air for heating and cooling. Ambient air is drawn into the machine by a small fan and warmed with a heating coil. Since air has a very low thermal capacity, the instrument can attain a thermal ramping rate of about 20°C per second.
Figure 1: Schematic drawing of the LightCycler® 2.0 Instrument.
PCR occurs in specially designed glass capillaries, which have an optimal surface-to-volume ratio to ensure rapid equilibration between the air and the reaction components. A typical amplification cycle requires only 30 to 60 seconds; an amplification reaction with 30 cycles is usually complete in about 30 minutes. To avoid contamination, the capillary is tightly closed; you do not need to open it at any time during analysis.
Two sizes of capillaries, 20 μl and 100 μl, are available. The standard 20 μl capillaries can be used with both the LightCycler® 1.5 (not available for sale) and 2.0 Instruments . The 100 μl size works only with the LightCycler® 2.0 Instrument and provides you more flexibility in designing your assay.
Up to 32 capillaries can be analyzed in a single run. The capillaries are placed in a sample carousel that rotates in the thermal chamber of the instrument. During each measurement, a stepper motor positions the sample carousel so the capillary tip is precisely at the focal point of the optical unit.
Fluorimeter and Light Source
A single microvolume fluorimeter is used to quantify amplification products. For fluorescence excitation, the LightCycler®2.0 Instrument uses a blue LED, which emits light at 470 nm. This LED, which is an integral part of the fluorimeter, needs no maintenance and has a long life span.
Both the high-quality optics, including the specialized light source and filters, and the glass capillaries, which focus most of the emitted light on the capillary tip, ensure that the measurement is extremely fast and sensitive. In a normal amplification run, measurement of a sample only takes about 20 milliseconds.
The LightCycler® 2.0 Instrument has a six-channel photometer that measure fluorescence at 530, 560, 610, 640, 670 and 705 nm: