VENTANA ANTI-MELANOMA PRIMARY ANTIBODY by VENTANA MEDICAL SYSTEMS, INC.

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 Description

AI/ML Overview

Here's an analysis of the provided text, focusing on the acceptance criteria and study information for the Ventana Medical Systems' Anti-Human Melanosome(Clone HMB45) device:

1. Table of Acceptance Criteria and Reported Device Performance

The original document does not explicitly state pre-defined acceptance criteria with numerical targets. Instead, it describes general desired performance characteristics (specificity, sensitivity, reproducibility) and then presents the results. Therefore, the "acceptance criteria" presented below are inferred from the desired characteristics mentioned and the "reported device performance" are the results provided.

Acceptance Criteria (Inferred)	Reported Device Performance
Specificity: No staining of normal cells.	Specific: No staining of normal cells (75 normal tissues). Confined to neoplastic melanocytic cells in adults, except for specific reactivities in "activated" melanocytes (junctional nevi, Spitz nevi, cellular blue nevi, junctional components of compound nevi) and very rarely in angiolipomas. `Note: The document highlights limitations where HMB45 is not useful for distinguishing benign from malignant proliferation due to reactivity with junctional nevi, Spitz tumors, and atypical melanocytic hyperplasia.`
Sensitivity: Consistent staining of melanoma tumors.	Sensitive: Consistent staining of 19 out of 20 melanoma tumors. This agrees with external reports (Gowan and associates: 60 of 62 melanomas positive). `Note: Sensitivity is stated to be dependent on tissue processing and slide preparation.`
Negative Control Performance: Negative results.	Negative Control: Negative results for negative controls run with each tissue.
Inter-run Reproducibility: Consistent positive staining and similar intensity across multiple runs.	Inter-run Reproducibility: 16 out of 16 samples of the same tissue stained positively across 16 different instrument runs. All slides had similar staining intensity.
Intra-run Reproducibility: Consistent positive staining and equivalent intensity within a single run.	Intra-run Reproducibility: 10 out of 10 samples of the same tissue stained positively within one run. All slides stained with equivalent staining intensity.
Appropriate Staining: Staining only in abnormal cells of melanocytic lineage, not inappropriate tissues.	Appropriate Staining: Staining occurred in the cytoplasm of melanoma cells. No inappropriate staining of tissues in the study. Does not react with non-melanocytic tissues (epithelial, lymphoreticular, glial, mesenchymal origin), with rare exceptions like myoid component of RAML and angiomyolipoma. `Note: The document mentions early evidence of unexpected reactivity in RAML and angiomyolipoma, which seem like potential "inappropriate" staining but are described as related antigen or regular expression.`

2. Sample Size Used for the Test Set and Data Provenance

Test Set Sample Size:
- Specificity: 75 normal tissues, an unspecified number of "melanoma tumors" for the initial specificity check, and an unspecified number of "pathologic samples" in the comparative study.
- Sensitivity: 20 melanoma tumors.
- Reproducibility: 16 samples (for inter-run), 10 samples (for intra-run).
- Total for overall comparative study: "Formalin fixed paraffin embedded preparations from normal and pathologic samples". The exact count is not provided for the full comparative study beyond the specific breakdowns for specificity, sensitivity, and reproducibility.
Data Provenance: Retrospective. Samples were "obtained from excess tissues obtained for reasons other than the present study." The country of origin is not specified.

3. Number of Experts Used to Establish the Ground Truth for the Test Set and Their Qualifications

Number of Experts: One ("a qualified pathologist").
Qualifications: "a qualified pathologist". No further specific details (e.g., years of experience, subspecialty) are provided.

4. Adjudication Method for the Test Set

Adjudication Method: None explicitly stated as a multi-reader consensus process. The evaluation was performed by "a qualified pathologist." It appears to be a single-reader assessment.

5. If a Multi-Reader Multi-Case (MRMC) Comparative Effectiveness Study Was Done

No, a multi-reader multi-case (MRMC) comparative effectiveness study was not explicitly described. The study focused on the performance of the antibody in staining tissues, evaluated by a single pathologist. There is no mention of human readers improving with or without AI assistance, as AI is not a component of this device.

6. If a Standalone (i.e., algorithm only without human-in-the-loop performance) Was Done

This question is not applicable. The device is an immunohistochemical reagent (an antibody), not an algorithm or AI system. Its performance is inherently "standalone" in the sense that it's the chemical's reaction, which is then interpreted by a human.

7. The Type of Ground Truth Used

Type of Ground Truth: Pathological diagnosis. The samples were "pathologic and normal tissues." The evaluation was done by a "qualified pathologist" who assessed "specific staining intensity and background staining." The inherent assumption is that the pre-existing diagnosis of "melanoma" or "normal tissue" served as the ground truth against which the antibody's staining was compared.

8. The Sample Size for the Training Set

This device is an antibody, not a machine learning model, so there is no "training set." The antibody was developed through a process involving immunization and hybridoma screening.
For the development of the antibody: The immunogen was "Melanoma metastases from axillary lymph node finely minced in phosphate buffered saline." Hybridomas were screened by immunocytochemistry on "Carnoy's fixed, paraffin-embedded sections of the same melanoma that was used as an immunogen." The exact number of screens/samples used in this development phase (analogous to a training set for an AI) is not specified.

9. How the Ground Truth for the Training Set Was Established

This question is not applicable in the context of an antibody's development as an AI training set.
For the development of the antibody: The "ground truth" during development was the ability of the antibody to react with the target antigen in melanoma cells, specifically the "oncofetal antigen present within immature melanosomes." This was confirmed by the screening process using the same melanoma tissue that served as the immunogen, indicating a direct confirmation of its binding to the intended target.

Summary

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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 melanosomes . 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 intraperitoreally 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 immunogen´.

In normal tissues. HM845 has been shown to react with immature melanosomes in fetal and neonatal melanocytes and infantile retinal pigment epithelium'-. It does not react with normal resting adult melanocytes, regardless of the degree of pigmentation 3.0 The presence of the antigen indicates active melanosome
formation and thus melanocytic differentiation? - 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 grandation tjssue, hemangionas, vessel -rich tumor stroma, and basa1 cell
carcinomas: 3.14. - Occasional pigmented cells of hair follicles have beerved with melanocytes in lentigines or overlying fibroblastic proliferations, such as keloids, dermatofibromas and old fibrotic hemangiomas . HMB45 does not appear to react with any non-melanocytic tissues, and its expression in adults is confined to neoplastic melanocytic cells'.

Many fetal antigens are re-expressed in oncogenic tissue. In malignant cells,
HMB45 stains most primary and metastatic melanomas-3.4.5.10 . Several investigators have confirmed the high sensitivity and specificity of HMB45 for me1anoma3.4.s Desmoplastic malignant melanomas infrequently express the HMB45 antigen, possibly because they contain few melanosmes and have a mesenchymal
pattern of differentiation · Non melancytic tumor cells of epithelial, Tymphoreticular, glial and mesenchymal origin are not labeled; 13 lymphoreticular. glial and mesenchymal origin are not labeled: ". except for related antigen located in the myoid component of RAML appears to be similar to the HMB45 antigen present in melanomas 2

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 intrademal components of compound nevif 3.3.3. 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 hyperplasia´.

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. Samles 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 associates 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

Kapur RP, Bigler SA, Skelly M. Gown AM: Anti-melanoma monoclonal 1 . antibody HMB45 identifies an oncofeatal glycoconjugate associated with immature melanocytes. J Histochem Cytochem 1992;40:20/-212.
1. Gown AM, Vogel AM, Hoak D, Gough F, McNutt MA: Monoclonal antibodies specific for melanocytic tumors distinguish subpopulations of melanocytes. Am J Patho7 1986;123:195-203.
Colombari R. Bonetti F. Zamboni G. Scarpa A. Marino F. Tomezoli A.
Capelli P. Mestraina F. Chilosi M. Fiore-Donati L. Distribution of 3.

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

Ordonez NG, Xiaolong J. Hickey RC: Comparison of HMB45 monoclonal 4. antibody and S-100 protein in the immunohistochemical diagnosis of melanoma. Am J Clin Pathol 1988;90:385-390.
Wick MR, Swanson PE, Rocamora A: Recognition of malignant melanoma by ട്. monoclonal antibody HMB45. An immunohistochemical study of 200 paraffinembedded cutaneous tumors. J Cutan Pathol 1988;15:201-207.
Taylor RT and Cote RJ. Immunomicroscopy: A Diagnostic Tool for the 6. Surgical Pathologist. W.B. Saunders Company. Philadelphia, 1994.
1. 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.
Esclamado RM, Gown AM, Vogel AM. Unique proteins defined by monoclonal 8. antibodies specific for human melanoma. Am J Surg 1986:152:376-385.
Smoller BR, McNutt NS, Hsu A. HMB45 recognizes stimulated melanocytes. g . J. Cutan Pathol 1989; 16:49-53.
Smoller BR, Hsu A. Krueger J. HMB45 monoclonal antibody recognizes an 10. inducible and reversible melanocyte cytoplasmic protein. J Cutan Pathol 1991: 18:315-322.
Pea M. Bonetti F, Zamboni G, Martignoni G, Riva M, Colombari R, Mombello 11. 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.
Ashfaq R, Weinberg AG, Albores-Saavedra J. Renal angiomyolipomas and 12. HMB45 reactivity. Cancer 1993; 71:3091-3097.
Skelton HG. Smith KJ. Barrett TL. Lupton GP, Graham JH. HMB45 staining 13. in benign and malignant melanocytic lesions. Am J Derm 1991; 13:543-550.

Regulation Number and Section

§ 866.5550 Immunoglobulin (light chain specific) immunological test system.

(a)
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.(b)
Classification. Class II (performance standards).