Auto-Immunity & Ease of Use through Automation The DIAsource Neptune Instrument & Reagents
Technology used The test is based on the principle of an Enzyme Immunoassay. The test strips are composed of a membrane fixed on a specific plastic support. During the automated test procedure, the Neptune instrument sequentially incubates the strips in the wells of ready-to-use reagent cartridges. Briefly: 1. the strips are first incubated with diluted patients sera. Human antibodies, if present, bind to the corresponding specific antigen(s) on the membrane. 2. Unbound or excess antibodies are removed by washing. 3. Upon further incubation into AP-conjugated goat antibodies against human IgG, the enzyme conjugate binds to the antigen-antibody complexes. 4. After removal of excess conjugate by washing, the strips are finally incubated into a substrate solution. Enzyme activity, if present, leads to the development of purple dots on the membrane pads. The intensity of the coloration is directly proportional to the amount of antibody present in the sample.
DIAspot Neptune Characteristics Up to 12 auto-antibodies tested per strip Testing time = 64 minutes 2 built-in controls : o Reactive control or Positive control : validity of the test o Cut-off control or Negative control : interpretation of the results Qualitative (visual interpretation) or semiquantitative results (Quantification software) The reagents are provided in ready-to-use individual cartridges, to fit the Neptune automated instrument.
DIAspot Neptune Characteristics
DIAspot Neptune Characteristics 7/23/2015 5
DIAspot Neptune Characteristics Total processing time (1 24 samples ) = 64 min 7/23/2015 6
DIAspot Neptune Characteristics DIAspot Features Examples of interpretation
DIAspot Neptune Characteristics Maximum 12 auto-antibodies detected per test strip Only Qualitative/Semi-quantitative results Quantitation trough software is based on comparison of the intensity of each dot to that of the Positive Control (RC) and the Negative Control (Cut-off). The obtained intensity is expressed in Arbitrary Units (AU) on a scale from 0 (negative results) to 100 (1 100, results considered as positive)
2 DIAspot MultiQUANT Neptune KITS
DIAspot MultiQUANT Neptune characteristics First Microarrays in auto-immune diagnostics Up to 25 auto-antibodies detected and quantified per strip Fully automated in 64 min (Neptune Instrument) Results in triplicate : Calculation for each results of mean value and standard deviation
DIAspot MultQUANT Neptune characteristics (2) Many built-in controls : Reactive control or Positive control : validity of the test Sample control : must have a minimum pre-defined colour intensity Conjugate control : control of the conjugate used for the respective conjugate specific of the kit. Substrate kinetic : validity of the substrate 6 point calibration curve :
Many built-in controls : Sample control : must have a minimum pre-defined colour intensity 6 point calibration curve : 6 triplicate lines, including blank control of increasing colour intensity from top to bottom. Must fit a pre-determined specific curve equation. All 25 antigens are analysed in triplace : individual results and mean results available. 7/23/2015 12
DIAspot MultiQUANT Neptune characteristics (3) DIAspot MultiQUANT Features Example of Interpretation
3 NEPTUNE INSTRUMENT
General open systems inconvenients DOT s can be automated on open-systems (ex: Tecan, Medtec,...) BUT All these machines have pumps and tubes to manage reagents Pumps/tubes limit the possibility of using different reagents / kits in a same run lack of flexibility Sequential dispensing/filling operations create unequal incubation conditions from first to last well Drift phenomena Important dead volumes of liquids in the tubing often require expensive extra reagents, particularly when small series or single tests are regularly performed Increased test cost Necessity to clean and regularly replace the pumps and tubes time consuming and expensive, need for maintenance contracts No traceability of the test conditions problem for accredited laboratories
Neptune description (1) A No pumps, no tubes machine Each strip is provided on a strip holder Up to 24 strip holders can be loaded on a moving arm The moving arm moves from back to front and dives sequentially the strips in the wells of individual cartridges Each cartridge contains the ready-to-use reagents for a strip (no extra reagents or consumables!)
Neptune characteristics (1) Allows automation of DIAspot Neptune and DIAspot MultiQUANT Neptune strips more than 30 differents kits Up to 24 identical or completely different strips can be loaded on the moving arm. Classic Dot strips (DIAspot) can be mixed with Microarray strips (DIAspot MultiQUANT Neptune ) in the same run total flexibility Bar code system control the right association strip/cartridge traceability of the test No mismatch of reagents and tests
DIAspot Neptune characteristics (2) No priming, no tedious washing or disinfection steps, easy maintenance save on time No dead volume no extra volume of reagent to order allows small series or single tests to be run economically All the strips are loaded on the same moving arm Same incubation conditions for all the strips, no drift Small Instrument Space saving instrument No pumps, no valves, no liquid & tubing inside instrument No daily, weekly and monthly maintenance for lab technician No maintenance contract means no additional cost
Key Figures Neptune instrument Characteristics Weight Size Figures 9,5 kg 30x30x55cm Number of kits different per run Up to 24 Number of samples per run Up to 24 Volume of sample 10µl Preparation time Running time 15 minutes 45 minutes Maximum results per run 288 (DIAspot ) 600 (DIAspot MultiQUANT)
Summary (2) Characteristics Number of auto-antibodies detected per strip Testing DIAspot DIAspot Neptune DIAspot MultiQUANT Neptune Up to 12 Up to 12 Up to 25 Manual or Open systems Neptune Instrument Neptune Instrument Test duration 64 min 64min 64 min Dr SPOT software Accessory Accessory Compulsory Results Qualitative or Semiquantitative Qualitative or Semiquantitative Quantitative Built-in controls 2 2 3 and calibration curve
Neptune Quantification Software Version 3.4 7/23/2015 21
7/23/2015 Minimal system requirements of the computer The following system requirements are needed : Microsoft Windows 2000/Windows 2003/Windows 2008 server/xp/vista/windows 7 CD-ROM Driver 512Mb RAM 100 Mb free space on the hard disk USB receptacle Java Software installed Adobe Software installed These elements are not compulsory but are recommended: A printer to print out the working list and the results An internet connection to send the results by email/ A handheld barcode reader for the automatic entering of ID samples
7/23/2015 Specifications of the scanner The Neptune scanner has been designed for the digitalization of the images of 1 to 24 strips, mounted on a «Clamp». The Neptune scanner contains an integrated calibration curve for the grayscale which allows the calibration of the scanner at each reading.
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7/23/2015 The DIASpot Clamp : for the strips of the DIASpot kits. This clamp can be ordered from DIAsource. The Neptune Instrument Clamp : for the strips of the DIASpot Neptune and DIASpot Multi QUANT Neptune kits. This clamp is sent with the Neptune Instrument
7/23/2015 1. Description of the main commands. Command Working List
7/23/2015 The column Position indicates the position on the clamp (from 1 to 24) The column Kit indicates the type of kit which is used for the test. The column Lot indicates the lot used for the test. The column Strip Number indicates the strip s number The column ID sample contains the ID of the sample. The column Patient data contains information about the patient.
7/23/2015 DIASpot Neptune and/or DIASpot MultiQUANT Neptune strips have been performed on the Neptune Instrument and are therefore already mounted on the appropriate Neptune Instrument Clamp. Remove the Neptune Instrument Clamp from the Neptune Instrument, and let dry tested strips, the dots facing upwards (image 1). Turn the clamp so that the revealed dots face the Neptune scanner window and away from the operator (image 2). Finally, insert it into the lateral emplacement of the Neptune scanner (see image 3).
7/23/2015 Display of general results sheet