VENTANA ANTI-MELANOMA PRIMARY ANTIBODY

K941170 · Ventana Medical Systems, Inc. · DEH · Apr 22, 1997 · Immunology

Device Facts

• Record ID: K941170
• Device Name: VENTANA ANTI-MELANOMA PRIMARY ANTIBODY
• Applicant: Ventana Medical Systems, Inc.
• Product Code: DEH · Immunology
• Decision Date: Apr 22, 1997
• Decision: SESE
• Submission Type: Traditional
• Regulation: 21 CFR 866.5550
• Device Class: Class 2

Indications for Use

Ventana Medical Systems' Anti-Human Melanosome(Clone HMB45) may be used to aid in the identification of abnormal cells of melanocytic lineage as an aid in diagnosis of anaplastic tumors.

Device Story

Anti-Human Melanosome (Clone HMB45) is a mouse monoclonal antibody reagent used in immunohistochemistry. It binds to a 10kD neuraminidase-sensitive sialylated glycoconjugate in immature melanosomes. Used in clinical pathology laboratories, the reagent is applied to formalin-fixed, paraffin-embedded tissue sections, typically processed on automated slide stainers (e.g., Ventana ES). A pathologist evaluates the slides for cytoplasmic staining intensity and distribution. Positive staining indicates melanocytic differentiation, aiding in the identification of melanoma metastases and distinguishing melanocytic lineage in anaplastic tumors. The reagent assists in clinical decision-making by providing diagnostic confirmation of melanocytic origin, though it does not distinguish between benign and malignant melanocytic proliferations.

Clinical Evidence

Bench testing only. Study evaluated 75 normal tissues and 20 melanoma tumors. Sensitivity: 19/20 (95%) melanoma tumors stained positively. Specificity: 0/75 normal tissues stained. Inter-run reproducibility (16/16) and intra-run reproducibility (10/10) demonstrated consistent staining intensity using DAB detection kits.

Technological Characteristics

Mouse monoclonal antibody (Clone HMB45) targeting 10kD neuraminidase-sensitive sialylated glycoconjugate. Reagent for immunohistochemistry. Used with automated slide stainer (Ventana ES) and DAB detection kits. Formulated for formalin-fixed, paraffin-embedded tissue sections.

Indications for Use

Indicated for use as an aid in the diagnosis of anaplastic tumors by identifying abnormal cells of melanocytic lineage in tissue samples.

Regulatory Classification

Identification

An immunoglobulin (light chain specific) immunological test system is a device that consists of the reagents used to measure by immunochemical techniques both kappa and lambda types of light chain portions of immunoglobulin molecules in serum, other body fluids, and tissues. In some disease states, an excess of light chains are produced by the antibody-forming cells. These free light chains, unassociated with gamma globulin molecules, can be found in a patient's body fluids and tissues. Measurement of the various amounts of the different types of light chains aids in the diagnosis of multiple myeloma (cancer of antibody-forming cells), lymphocytic neoplasms (cancer of lymphoid tissue), Waldenstrom's macroglobulinemia (increased production of large immunoglobulins), and connective tissue diseases such as rheumatoid arthritis or systemic lupus erythematosus.

Submission Summary (Full Text)

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K941170
APR 22 1997

# 510(k) Summary of Safety and Effectiveness

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.

Ventana Medical Systems' Anti-Human Melanosome(Clone HMB45) may be used to aid in the identification of abnormal cells of melanocytic lineage as an aid in diagnosis of anaplastic tumors. Anti-Human Melanosome(Clone HMB45) specifically binds to an oncofetal antigen present within immature melanosomes in the cytoplasm of melanoma cells and prenatal and infantile epithelium. This product is substantially equivalent to the same clone sold by a different manufacturer.

Ventana Medical Systems' Anti-Human Melanosome(Clone HMB45) contains a mouse monoclonal antibody directed against a 10kD neuraminidase sensitive sialylated glycoconjugate present in immature melanosomes¹. Melanoma metastases from axillary lymph node finely minced in phosphate buffered saline was used as the immunogen. The spleen from a BALB/c mouse injected intraperitoneally with the minced tissue was fused with NS-1 cells. Hybridomas were screened by immunocytochemistry on Carnoy's fixed, paraffin-embedded sections of the same melanoma that was used as an immunogen².

In normal tissues, HMB45 has been shown to react with immature melanosomes in fetal and neonatal melanocytes and infantile retinal pigment epithelium¹,². It does not react with normal resting adult melanocytes, regardless of the degree of pigmentation²,³,⁸. The presence of the antigen indicates active melanosome formation and thus melanocytic differentiation⁹,¹³. Adult melanocytes are capable of re-expressing the fetal antigen upon activation. Melanocytes activated by a variety of stimuli have been observed. For example, HMB45 positive cells have been detected in tissue overlying or adjacent to granulation tissue, hemangiomas, vessel-rich tumor stroma, and basal cell carcinomas³,⁹,¹⁰,¹³. Occasional pigmented cells of hair follicles have been observed to stain positively². HMB45 staining has not been observed with melanocytes in lentigines or overlying fibroblastic proliferations, such as keloids, dermatofibromas and old fibrotic hemangiomas⁹. HMB45 does not appear to react with any non-melanocytic tissues, and its expression in adults is confined to neoplastic melanocytic cells¹.

Many fetal antigens are re-expressed in oncogenic tissue. In malignant cells, HMB45 stains most primary and metastatic melanomas²,³,⁴,⁵,⁸,¹³. Several investigators have confirmed the high sensitivity and specificity of HMB45 for melanoma³,⁴,⁵. Desmoplastic malignant melanomas infrequently express the HMB45 antigen, possibly because they contain few melanosomes and have a mesenchymal pattern of differentiation²,¹³. Non melanocytic tumor cells of epithelial, lymphoreticular, glial and mesenchymal origin are not labeled¹,³,⁵ except for renal angiomyolipomas (RAML), a type of mesenchymal hamartoma¹¹,¹². An HMB45-related antigen located in the myoid component of RAML appears to be similar to the HMB45 antigen present in melanomas¹².

HMB45 is reactive with "activated" melanocytes as seen in melanomas, junctional nevi, junctional components of compound nevi, Spitz nevi, and cellular blue nevi, while it is nonreactive with common acquired and intradermal nevi and intradermal components of compound nevi²,³,⁶,¹³. Because of this differential staining on pigmented cells, HMB45 immunostaining may be a

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useful adjunct for measuring the Breslow depth of melanomas, since it is positive on melanoma cells and negative on deep nevi cells that may accompany melanomas⁶,⁷. Unfortunately, HMB45 is not useful in distinguishing benign and malignant melanocytic proliferation because it recognizes junctional nevi. Spitz tumors and atypical melanocytic hyperplasia².

## Comparative Study

Supporting data for the equivalence statement is shown by the following study. Formalin fixed paraffin embedded preparations from normal and pathologic samples were tested using the Ventana Anti-Human Melanosome Antibody. Samples were obtained from excess tissues obtained for reasons other than the present study. Pathologic and normal tissues were examined. Slides were processed on the Ventana ES Automated Slide Stainer, prepared for examination, and evaluated by a qualified pathologist for specific staining intensity and background staining.

## Results

Staining occurred in the cytoplasm of melanoma cells. Negative control tissue was all negative. There was no inappropriate staining of the tissues in this study.

Specificity of the antibody was shown by no staining of normal cells of 75 normal tissues and staining of cells of melanoma tumors.

The sensitivity of this antibody was shown by consistent staining of 19 of 20 melanoma tumors. This agrees with the report Gowan and associates² where 60 of 62 melanomas showed positive staining. As with any immunohistochemical reagent, the sensitivity is dependent on tissue processing and slide preparation parameters. The negative control which was run with each tissue gave negative results.

Inter-run reproducibility was determined based on samples of the same tissue on 16 different instrument runs using the antibody and DAB detection kits. Sixteen of 16 stained positively. All slides has similar staining intensity. Intra-run reproducibility was determined based on 10 samples of the same tissue within one run using the antibody and DAB detection kits. Ten of 10 slides stained positively. All slides stained with equivalent staining intensity.

## References

1. Kapur RP, Bigler SA, Skelly M, Gown AM: Anti-melanoma monoclonal antibody HMB45 identifies an oncofeatal glycoconjugate associated with immature melanocytes. J Histochem Cytochem 1992;40:207-212.
2. Gown AM, Vogel AM, Hoak D, Gough F, McNutt MA: Monoclonal antibodies specific for melanocytic tumors distinguish subpopulations of melanocytes. Am J Pathol 1986;123:195-203.
3. Colombari R, Bonetti F, Zamboni G, Scarpa A, Marino F, Tomezzoli A, Capelli P, Mestraina F, Chilosi M, Fiore-Donati L: Distribution of

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melanoma specific antibody (HMB45) in benign and malignant melanocytic tumors. Virchows Archiv A Pathol Anat 1988;413:17-24.

4. Ordonez NG, Xiaolong J, Hickey RC: Comparison of HMB45 monoclonal antibody and S-100 protein in the immunohistochemical diagnosis of melanoma. Am J Clin Pathol 1988;90:385-390.

5. Wick MR, Swanson PE, Rocamora A: Recognition of malignant melanoma by monoclonal antibody HMB45. An immunohistochemical study of 200 paraffin-embedded cutaneous tumors. J Cutan Pathol 1988;15:201-207.

6. Taylor RT and Cote RJ. Immunomicroscopy: A Diagnostic Tool for the Surgical Pathologist. W.B. Saunders Company. Philadelphia, 1994.

7. Duray PH, Palazzo J, Gown AM, Ohuchi N. Melanoma cell heterogeneity. A study of two monoclonal antibodies compared with S-100 in paraffin sections. Cancer 1988; 61:2460-2468.

8. Esclamado RM, Gown AM, Vogel AM. Unique proteins defined by monoclonal antibodies specific for human melanoma. Am J Surg 1986;152:376-385.

9. Smoller BR, McNutt NS, Hsu A. HMB45 recognizes stimulated melanocytes. J. Cutan Pathol 1989; 16:49-53.

10. Smoller BR, Hsu A, Krueger J. HMB45 monoclonal antibody recognizes an inducible and reversible melanocyte cytoplasmic protein. J Cutan Pathol 1991; 18:315-322.

11. Pea M, Bonetti F, Zamboni G, Martignoni G, Riva M, Colombari R, Mombello A, Bonzanini M, Scarpa A, Ghimenton C, Donati LF. Melanocyte-marker-HMB45 is regularly expressed in angiomyolipoma of the kidney. Pathol 1991; 23:185-188.

12. Ashfaq R, Weinberg AG, Albores-Saavedra J. Renal angiomyolipomas and HMB45 reactivity. Cancer 1993; 71:3091-3097.

13. Skelton HG, Smith KJ, Barrett TL, Lupton GP, Graham JH. HMB45 staining in benign and malignant melanocytic lesions. Am J Derm 1991; 13:543-550.