Molecular HLA Typing
Immucor Lifecodes HLA Class I & Class II SSO Typing
Inno-Train HLA Ready Gene PCR-SSP
Inno-Train HLA Ready Gene for Disease-Associated Typing
Inno-Train Ready Gene Wipe Test
Inno-Train HLA FluoGene
Inno-Train SuBiTo HLA Sequence-Based Typing
Immucor Lifecodes HLA products are based on Luminex xMAP technology. The Lifecodes HLA-SSO Typing kits utilise sequence-specific oligonucleotides (SSOs) to identify which human leucocyte antigen (HLA) alleles are present in a PCR-amplified sample. It is the set of SSOs employed that determines the ability to distinguish among the various alleles present in the PCR amplification. All assays are carried out in a single reaction vessel with the addition of a single hybridization reagent for each locus tested.
Luminex Key Features:
- Amplified DNA can be analysed with up to 100 different SSO probes simultaneously
- Multiplex reactions reduce labour, consumables and assay time
- XY platform allows walk-away acquisition from 1 to 96 samples at a time
- Convenience of full automation, if required, through collaboration with Hamilton Robotics
Lifecodes HLA SSO Typing Kits Key Features:
- Unique probe design minimises ambiguities
- Master mix includes primers, dNTPs and PCR buffer, eliminating extra steps and consumables
- Homogeneous assay eliminates the need for centrifugation and wash steps, saving time and reducing the chance of pipetting errors
- Analysis software automatically imports data from the Luminex instrument
HLA Ready Gene is a molecular biological product line for detection of blood group systems based on PCR-SSP, which presents the ideal complement to serological blood group typing. The modular kit design gives the freedom to choose which systems to use for maximum flexibility.
Inno-Train’s molecular SSP detection systems are based on the Polymerase Chain Reaction (PCR), which enables amplification of defined DNA sequences. After successful amplification, the genomic DNA target sequence is present in a detectable concentration. The term Sequence Specific Priming (SSP), describes a variant of PCR in which only the sequence of the primer at the 3‘ end is responsible for specification of the allele to be identified. For complete PCR SSP analysis, several amplifications are performed in parallel. Evaluation of the result is performed by agarose gel electrophoresis. Under UV light, the bands are visualised by intercalating ethidium bromide and can be photographed for documentation.
HLA Ready Gene kits for disease-association typing are based on the SSP technique. The primer mixes are pre-aliquoted and dried, and all kits include negative control and ReadyPCR Mix, containing cresol red, glycerine, dNTPs and PCR buffer. HLA Ready Gene kits include:
- HLA Ready Gene B27 Low
- HLA-Ready Gene B5/57 cross
- HLA-Ready Gene Coeliac Disease
- HLA-Ready Gene B57 Low
- HLA-Ready Gene Narcolepsy
Inno-Train’s Ready Gene Wipe Test enables the user to monitor for contamination with amplicons created by the Ready Gene product line on working areas. It can also be used as a contamination control in Inno-Train’s Ready Gene typing kits.
The primer mix contains the primer pairs that amplify the 430 bp HGH fragment (human growth hormone gene), which is also part of all internal control amplicons in Inno-Train’s Ready Gene typing kits.
FluoGene is a new typing method which combines the speed of the PCR-SSP method with Endpoint Fluorescence detection, without the need for agarose gel electrophoresis. FluoGene can be automated easily and flexibly: in its basic configuration the system uses the FluoVista Analyser in addition to a standard thermal cycle and the level of automation can be increased by adding a small pipetting station or a complete liquid handling processor with integrated modules.
Benefits of the FluoGene System:
- Same system for HLA, HPA and RBC
- Minimal hands-on time
- All reagents are included and pre-aliquoted
- No gel electrophoresis
- Only 7.5ng DNA required per reaction
- No risk of contamination
- Easy software based analysis
- Automation available
The SuBiTo system from Inno-Train is for high resolution HLA typing by DNA sequencing. After amplification of the HLA genes by PCR, the HLA alleles present are typed using a DNA sequencer. All systems have been validated using an ABI capillary sequencer. The method is based on “Sanger“ sequencing with fluorescent-marked dideoxynucleotides.
The complete SuBiTo sequencing method requires genomic DNA as its basic material and is divided into 5 focal working steps:
- PCR amplification of genomic HLA sequences
- Enzymatic purification of the PCR products
- Sequencing reactions (Cycle Sequencing)
- Purification of the sequencing products, precipitation
- Sequencing (automated), detection and analysis of the DNA chains