VARIANT HEMOGLOBINOPATHY PROGRAM

K954048 · Bio-Rad · GKA · Nov 20, 1995 · Hematology

Device Facts

Record IDK954048
Device NameVARIANT HEMOGLOBINOPATHY PROGRAM
ApplicantBio-Rad
Product CodeGKA · Hematology
Decision DateNov 20, 1995
DecisionSESE
Submission TypeTraditional
Regulation21 CFR 864.7415
Device ClassClass 2

Indications for Use

An aid in the detection of normal and abnormal hemoglobins by High Pressure Liquid Chromatography.

Device Story

The Variant Hemoglobinopathy Program is an automated HPLC system for hemoglobin analysis. It processes whole blood samples mixed with hemolysis reagent. The system uses a cation exchange cartridge and a linear buffer gradient to separate hemoglobin variants. A dual-wavelength photometer (415 nm and 690 nm) monitors elution; the 690 nm filter corrects baseline effects from buffer ionic strength changes. The system compares sample retention times against a Retention Time Marker containing known hemoglobins (D, S, C, E/A2, A2, F) to determine the hemoglobin phenotype. Used in clinical laboratory settings, the system automates the separation and identification process, replacing manual visual inspection of electrophoresis bands. This provides objective, reproducible identification of hemoglobin variants, aiding clinicians in diagnosing hemoglobinopathies.

Clinical Evidence

Accuracy was established via a correlation study comparing the Variant Hemoglobinopathy Program to the Isolab RESOLVE®-Hb test using 119 clinical samples. The study showed 100% agreement across 11 distinct hemoglobin phenotypes. Precision was evaluated per NCCLS EPS-T2 guidelines; within-run %CV ranged from 0.2% to 0.3%, and total %CV ranged from 0.4% to 0.9% for various hemoglobins (F, A2, E, D, S, C). No clinical prospective trials were required; evidence is based on analytical performance and method comparison.

Technological Characteristics

Cation exchange high-performance liquid chromatography (HPLC) system. Utilizes a cation exchange cartridge, elution buffers, and a dual-wavelength photometer (415/690 nm). Operates at 25 °C. Automated sample injection via auto-sampler. Requires 5 μl whole blood sample volume. Software-based retention time analysis for phenotype identification.

Indications for Use

Indicated for the detection of normal and abnormal hemoglobins in human blood samples to assist in the diagnosis of hemoglobinopathies.

Regulatory Classification

Identification

An abnormal hemoglobin assay is a device consisting of the reagents, apparatus, instrumentation, and controls necessary to isolate and identify abnormal genetically determined hemoglobin types.

Special Controls

*Classification.* Class II (special controls). A control intended for use with an abnormal hemoglobin assay is exempt from the premarket notification procedures in subpart E of part 807 of this chapter subject to the limitations in § 864.9.

Predicate Devices

Submission Summary (Full Text)

{0} BIO-RAD Bio-Rad Laboratories Diagnostics Group 4000 Alfred Nobel Drive Hercules, California 94547 Telephone: 510724-7000 Fax: 510741-5824 # SUMMARY OF SAFETY AND EFFECTIVENESS Submitter: Bio-Rad Laboratories, Inc. Clinical Systems Division 4000 Alfred Nobel Drive Hercules, California 94547 Phone 1-510-741-6015 FAX 1-510724-5024 K840018 Contact Person: John W. Nelson Manager, Regulatory Affairs NOV 20 1995 Date Prepared: Product Trade Name: Variant Hemoglobinopathy Program Common Name Variant Hemoglobin Testing System Classification Name: Abnormal Hemoglobin Assay, 81GKA The Variant Hemoglobinopathy Program is designed for use on the fully automated Variant analyzer. The analytical system consisting of instrument and reagent kit provides an assay for the detection of normal and abnormal hemoglobins by High Performance Liquid Chromatography, HPLC. To establish substantial equivalence to an existing device, and thus establish the safety and effectiveness of the Variant hemoglobinopathy Program, the hemoglobinopathy program has been compared to the Isolab RESOLVE®-Hb test (K840011). A review of the intended use of each system shows them to be essentially the same. The intended use of the Variant Hemoglobinopathy Program is stated as: An aid in the detection of normal and abnormal hemoglobins by High Pressure Liquid Chromatography. The intended use of the Isolab RESOLVE®-Hb test is states as: Designed for the separation of hemoglobin variants in adult and neonatal blood hemolysates by isoelectric focusing electrophoresis. The Variant Hemoglobinopathy Program utilizes the principles of cation exchange high performance liquid chromatography (HPLC). A hemolysate is prepared by mixing whole blood with a hemolysis reagent included in the kit. The prepared hemolysate is placed into the instrument's auto sampler. The sampler injects a specified amount of hemolysate onto the cartridge. The separation of normal and abnormal hemoglobins is accomplished on the cation exchange cartridge using a linear buffer gradient. A dual wavelength photometer (415 and 690 nm) monitors the elution of the separated hemoglobins from the cartridge, detecting absorbance changes at 415 nm. The 690 nm secondary filter corrects the baseline for effects caused by the mixing of buffers of different ionic strengths as the gradient is formed. {1} Retention time windows (ranges) are established for the presumptive identification of normal and abnormal hemoglobin based on their characteristic retention times. The Variant Hemoglobinopathy Program utilizes a Retention Time Marker containing hemoglobins D, S, C, E/A₂, A₂ and F. The initial retention times for the Variant Hemoglobinopathy Program Retention Time Marker were established using patient samples with confirmed normal and abnormal hemoglobins. In an assay, the retention times of the hemoglobins in the marker are compared to the retention times of the hemoglobins in a sample to determine the hemoglobin phenotype of the individual from whom the sample was obtained. The Isolab RESOLVE®-Hb test utilizes isoelectric focusing (IEF) to separate the hemoglobins in a prepared hemolysate. Mixtures of low molecular weight amphoteric molecules with varying pi's are contained in an agarose gel matrix an generate a stable pH gradient when an electric field is applied. When a sample is placed in this pH gradient generated by an electric field, it will migrate towards the anode or cathode until it reaches its isoelectric point. Once this happens, diffusion is counteracted by the electric field, and hemoglobin variants appear as discreet bands at their different isoelectric points. After separation is complete the bands are fixed, stained and compared by "eye" to known hemoglobins run under the same conditions for identification. The performance of the Variant Hemoglobinopathy Program was evaluated for precision, accuracy, and limits of detection. The precision studies were done according to NCCLS Vol. 12, No. 4, EPS-T2,p31, and are summarized below in Table 1. Accuracy was determined by a correlation study against the Isolab RESOLVE®-Hb test the results of which are shown below in Table 2. Table 3 compares the general characteristics of the two methods. Table 1 Precision Study | Sample Hemoglobin | Mean Retention Time (min.) | Within-Run %CV | Total %CV | | --- | --- | --- | --- | | F | 6.32 | 0.3 | 0.9 | | A₂ | 9.76 | 0.2 | 0.7 | | A₂ | 10.55 | 0.2 | 0.7 | | E | 10.43 | 0.2 | 0.6 | | D | 11.60 | 0.2 | 0.4 | | S | 12.00 | 0.2 | 0.4 | | C | 15.55 | 0.2 | 0.4 | {2} Table 2 Accuracy Study | Hemoglobin Phenotype | N | Hemoglobinopathy Program | | Isolab Resolve | | | --- | --- | --- | --- | --- | --- | | | | + | . | + | | | AA | 71 | 71 | 0 | 71 | 0 | | AS | 14 | 14 | 0 | 14 | 0 | | AC | 13 | 13 | 0 | 13 | 0 | | AE | 7 | 7 | 0 | 7 | 0 | | EE | 3 | 3 | 0 | 3 | 0 | | SS | 4 | 4 | 0 | 4 | 0 | | SC | 2 | 2 | 0 | 2 | 0 | | AD | 2 | 2 | 0 | 2 | 0 | | DS | 1 | 1 | 0 | 1 | 0 | | AzAz | 1 | 1 | 0 | 1 | 0 | | ACGPhiladelphia | 1 | 1 | 0 | 1 | 0 | | Total | 119 | 119 | 0 | 119 | 0 | Note: "+" denotes a positive identification, "-" denotes no hemoglobin detected. Table 3 Comparison of General Characteristics | Comparison of Methods | | | | --- | --- | --- | | Parameter | Variant Hemoglobinopathy Program | Isolab RESOLVE-Hb | | Analytes | Normal and abnormal hemoglobins | Normal and abnormal hemoglobins | | Measurement | Qualitative | Qualitative or quantitative using a densitometer | | Basic Principle | Charge based separation using liquid chromatography | Charge based separation using isoelectric focusing electrophoresis | | Instrumentation | Variant Hemoglobin Testing System | Isoelectric focusing electrophoresis unit | | Analysis Medium | Cation exchange high performance liquid chromatography | Agarose gel electrophoresis | | Temperature | 25 °C | 15 °C | {3} | Compariso_{n} of Methods (Continued) | | | | --- | --- | --- | | Parameter | Variant Hemoglobinopathy Program | Isolab RESOLVE-Hb | | Sample | Whole Blood | Whole Blood | | Sample Volume | 5 μl | 25 μl | | Reagents | Elution Buffers (1&2), Analytical cartridge (column), Retention Time Marker/Diluent, Hemolysis Reagent | IEF Gels, Anode Solution, Cathode Solution, Sample Prep Reagent, IEF Electrode Wicks, Sample Application Template, Blotting Paper | It can be concluded from the correlation study between the Bio-Rad Variant Hemoglobinopathy Program and the Isolab RESOLVE®-Hb test, and the similarities of the general characteristics, that the two assays are substantially equivalent. Based on the establishment of substantial equivalence, the safety and effectiveness of the Bio-Rad Variant Hemoglobinopathy Program is confirmed.
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