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Background Low-risk stage IB/IIA cutaneous melanoma represents an unmet need: a sizable population remains at clinically relevant risk of recurrence and death while adjuvant immunotherapy is approved only for stages IIB-IV. The previously clinically validated Immunoprint® 8-slide/7-biomarker IHC assay (7-P-IHC) robustly stratifies these ‘hidden high-risk’ patients. To improve laboratory throughput and enable single-platform commercialization, we bridged the assay to a 2-slide/7-biomarker multiplex immunofluorescence (mIF) format running on the Akoya PhenoImager HT. Here, we report analytical bridging data comparing mIF with the legacy IHC assay, and the mIF assay performance.Methods The 7-P-IHC Immunoprint® assay was adapted into a 2-slide/7-biomarker Opal™ mIF assay, leveraging the PhenoImager HT imaging system. Formalin-fixed, paraffin-embedded (FFPE) tissue from n = 53 primary melanomas (6 IA, 18 IB, 15 IIA, 14 IIB) were stained with multiplex Opal™ assay and original IHC assay. Pre-specified success criteria were overall/positive/negative percentage agreement (OPA/PPA/NPA) >85%, Cohen’s κ ≥ 0.60 for each biomarker and Pearson r ≥ 0.75 for the composite risk score. mIF panel staining, image acquisition and image unmixing were automated; risk scores were calculated blinded to the reference 7-P-IHC result. Panel performance was evaluated through equivalency between ‘gold-standard’ IHC and mIF, evaluation of umbrella effect, spectral crosstalk through drop test, and intra- and inter-day triplicate precision studies for assay reproducibility.Results The mIF assay met all pre-specified criteria with OPA = 96.2% (95% CI 87.0–99.0), PPA = 100% (86.7–100) and NPA = 88.9% (65.3–98.6). Strong analytical agreement was observed between all seven individual biomarker scores (Cohen’s κ 0.61–0.81), and for the composite risk score (Pearson r = 0.84). Strong equivalency was observed between IHC and mIF panels. No umbrella effect or spectral crosstalk was observed in drop test. High reproducibility was observed in inter-day/intra-day precision study and passed the preset criteria by board-certified pathologists.Conclusions The 2-slide/7-biomarker mIF Immunoprint® assay demonstrates analytical equivalence to the clinically validated 7-P-IHC assay in this analytical-bridging pilot and is not powered for outcome inference. These data satisfy CLSI EP09-A2 guidance for platform, and will be supplemented by an independent verification cohort to support forthcoming clinical-utility studies.Ethics Approval All FFPE blocks are commercially purchased from Precision for Medicine, IRB review Protocol Precision for Medicine - PFM004, REMNANT BIOSPECIMEN PROGRAM (Pro00051469). Advarra IRB reviewed the protocol referenced above, the Consent templates for the study and other supporting information.