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Questions regarding the OSNA® method

What does the abbreviation OSNA mean?

OSNA® stands for »one-step nucleic acid amplification.« The method involves the homogenisation of lymph node tissue followed by analysis of a molecular target mRNA called CK19 by the process of »reverse-transcription-loop-mediated-isothermal-amplification« (RT-LAMP). The name OSNA® is proprietary by Sysmex.

How does the LAMP-reaction work in principle?

LAMP which stands for Loop-mediated Isothermal Amplification is a simple, rapid and specific nucleic acid amplification method solely developed by Eiken Chemical Co., Ltd. It is characterised by the use of 6 different primers specifically designed to recognize 6 or 8 distinct regions on the target sequence. The reaction utilises a DNA Polymerase with strand displacement activity and proceeds at a constant temperature of around 65°C. Amplification and detection of the target sequence can be completed in a single step.

What are the characteristics of OSNA®?

The method is characterised by

  • No requirement for an RNA purification step.
  • A short reaction time consistent with the intra-operative use of the system.
  • The use of 6 primers that lead to a high specificity for the target mRNA.
  • Optimal Assay Design avoiding the amplification of CK19 Pseudogenes.
  • Gene-amplification at a constant temperature of 65°C to avoid genomic DNA amplification.

How long does an amplification reaction take on the RD100i Robotic Analyser System?

It takes 16 minutes.

How fast is the whole procedure of a sample analysis?

It takes about 30 minutes including pre-treatment for one lymph node and 40 minutes for 4 lymph nodes.

How many lymph nodes can be analyzed simultaneously?

Up to 4 lymph nodes can be processed in one run.

Can remaining lymph node lysate be stored?

The lysate can either be stored at -80°C or in liquid nitrogen.

Does the blue dye interfere with the OSNA® reaction?

The blue dye used in the surgical identification of Sentinel Lymph Nodes does not have an impact on the OSNA® result.

Questions regarding the reagents

What is the storage temperature of the reagents (Lynorhag and Lynoamp)?

Please store the homogenising reagent (Lynorhag) at 4°C and the amplification kit (Lynoamp) at -20°C.

How often can I thaw and freeze primers, controls and calibrator solutions?

The solutions may be freeze thawed upto 5 times.

How long can the reagents be kept outside the freezer – either on ice or in the instrument?

LynoAmp reagents can be kept on ice or on board the RD100i Robotic System for about 6 hrs.

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