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Vaccines, Blood & Biologics

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BK030056 510K Summary

This summary of 510(k) safety and effectiveness  information is being submitted in accordance with the requirements of SMDA 1990 and 21 CFR 807.92.

The assigned 51O(k) number is:               _,_

Submitter:      Dynal Biotech Ltd, UK

Address:                    11  Bassendale Road, Croft Business Park, Bromborough,
Wirral CH62 3QL, UK

Phone: Fax:

+44 151 346 1234
+44 151 346 1223
Contact:         Peter Maguire, Vice President Dynal Biotech, HLA Diagnostics

Summary Report Date: 11th August 2003

1.0    Introduction

The HLA loci of the human Major Histocompatibility  Complex  (MHC) located on the short arm of human chromosome 6 encode two distinct classes of highly polymorphic  cell surface molecules that bind and present processed  antigens in the form of peptides to T lymphocytes.  This presentation  step is crucial in initiating both cellular and humoral immune responses; consequently these molecules  play a central role in the regulation of the immune system, in transplantation  biology, as well as in susceptibility to a number of diseases, including auto-immune disordersand certain cancers.

The class I molecules,  HLA-A, HLA-B, and HLA-C, are found on most nucleated cells.  They are cell surface glycoprotein's that bind and present processed peptides derived from endogenously  synthesized proteins.(e.g., viral and tumour­ peptides) to CDB+T cells.  These heterodimers consist of an HLA-encoded  alpha chain associated with the non-MHC-encoded polypeptide, 132-microglobulin.

While 132-microglobulin is monomorphic, the alpha-chain genes are extremely polymorphic. This variability is localized primarily in exons 2 and 3, which encode the amino-terminal extracellular domains that function as the peptide-binding site. Within these two exons, the polymorphism  is concentrated  into discrete clusters that lie within a relatively conserved framework region. Analysis of HLA class I crystal structures, has shown that these polymorphic residues line the peptide­ binding cleft and interact directly with peptide and/or the T-cell receptor.

2.0    In Vitro Diagnostic Product Name

Proprietary Name:              Dynat RELl™ SSO HLA-ATyping Kit

 Common Name: HLA-A Typing Kit

Classification Name:            Unknown

3.0    Establishment Registration Number

Device Establishment Registration Number:         To be Issued

4.0     Device Classification

HLA Typing Reagents have been classified as Class I devices

5.0    Compliance to Classification Requirements

No performance standards have been established for HLA typing reagents. Dynal Biotech intends to comply with any standards applicable to the product developed in the future.
 

6.0    Intended Use

Dynal RELI™ SSO HLA-A typing kit provides a low to medium resolution HLA-A
typing result.

7.0    Device Description

The Dynal RELI™ SSO HLA-A Test is based on three major processes: PCR target amplification, hybridisation of the amplified products to an array of immobilized  sequence-specific oligonucleotide probes, and detection of the probe-bound amplified product by colour formation.

PCR Amplification  Reaction

The PCR reagent mixture containing the DNA specimen is heated to 95°C, separating the double-stranded DNA and exposing the specific primer target sequences. As the mixture cools, the biotinylated primers anneal to their targets. The thermostable recombinant  Thermus aquaticus (Taq) DNA polymerase in the presence of excess deoxynucleoside triphosphates (dNTPs), including deoxyadenosine, deoxyguanosine, deoxycytidine and deoxyuridine (in place of deoxythymidine), extends the annealed primers along the target templates to produce a biotinylated DNA sequence termed an amplicon. This process is repeated for a number of cycles, each cycle effectively doubling the amount of target DNA. For this test, the required number of cycles has been determined to be 35, theoretically yielding more than a billion-fold amplification.

Hybridisation  Reaction   _

After the PCR amplification process, the amplicons are chemically  denatured to form single-stranded DNA, these are added to a nylon membrane which contains an array of immobilized,  sequence-specific  oligonucleotide (SSO) probes. The biotin-labelled amplicons then bind (hybridise) to those SSO probes that contain
a complementary target sequence and thus are "captured" onto the membrane
strip.

A stringent wash step after hybridisation ensures the specificity of the reaction and removes all unbound amplicon.

Detection Reaction

The amplicon- probe complex is visualised using a colourmetric reaction. Streptavidin-horseradish peroxidase (SA-HRP) conjugate is added to the membrane and binds to the biotin-labelled amplicons captured by the SSO probe. Addition of hydrogen peroxide (H202)  and tetramethylbenzidine (TMB) substrate, results in the formation of a blue colour complex in the presence of SA-HRP. The resulting probe signals are compared to the control probe intensity and the samples hit pattern recorded for interpretation.

8.0    Proposed Labelling and Instructions for Use (IFU)

The proposed labels for Dynal RELI™ SSO HLA-A Typing Kit reagents are presented in Appendix A. The proposed IFU appears in Appendix B.

9.0    Statement of Substantial Equivalence

Summary of Substantial Equivalence

Dynal RELI™ SSO HLA-A Typing Kit provides a low to medium resolution HLA-A typing result.  It has been on sale for Research Only purposes since June 1999. Dynal Biotech now wishes to broaden the scope of this assay for clinical testing by gaining FDA clearance via this submission.

HLA-A typing at low to medium resolution is routinely performed worldwide using either a serological or DNA based technology. No single method was chosen to use as a predictive device to evaluate the performance of the RELI™ SSO typing system, as this reflects the diversity of commercial options available to customers and allows more customers to correlate performance data with the typing system they are currently performing.

Dynal RELI™ HLA-A Typing Kit is substantially equivalent to the following predictive devices

SSO: One Lambda Inc. LABType™ SSO (BK020055)
SSP: One Lambda Inc. MicroSSP™ (BK960062)
Serology: Biotest Diagnostics Lymphocyte HLA-AB 144 typing Tray
(BK830027).

Comparison of Dynal RELI™ HLA·A Typing Kit to Predictive Devices

All three predictive devices achieve a low to medium resolution HLA-A type with some differences in technology to Dynal RELI™ SSO. Dynal Biotech believes that RELI™ SSO maintains the same safety and effectiveness as the predictive devices.

RELI™ SSO:

The methodology is described in detail in section 2.0