Single-center ex-vivo study using blood samples from healthy donors processed through a 25-gauge vitrectomy system. The study compares four combinations of operational parameters (vacuum pressure and cut rate) to evaluate their impact on cellular recovery and morphological preservation. Pre- and post-procedure complete blood counts and cytology smears will be analyzed to identify the settings that minimize cell loss and mechanical damage, with the ultimate goal of optimizing diagnostic yield in vitreoretinal lymphoma (VRL) vitrectomy.
Primary vitreoretinal lymphoma (VRL) is a rare but aggressive intraocular malignancy, typically classified as a subtype of primary central nervous system lymphoma. It predominantly involves the retina and vitreous and is frequently associated with central nervous system disease. Clinical presentation often mimics chronic uveitis, leading to diagnostic delays and suboptimal management. Definitive diagnosis of VRL requires cytopathologic confirmation through diagnostic vitrectomy, which provides vitreous samples for cytology, immunohistochemistry, flow cytometry, and molecular analyses (e.g., MYD88 mutation testing, IL-10/IL-6 ratio). However, despite the central role of vitreous biopsy, its diagnostic sensitivity remains limited to approximately 70% (Iuliano L et al. Int Ophthalmol 43, 2841-2849 (2023)), primarily due to the mechanical fragility and scarcity of malignant lymphoid cells. During vitrectomy, the vitreous is fragmented and aspirated through a high-speed cutting probe (vitrector), which applies both mechanical cutting and vacuum forces. These operational parameters, while optimized for surgical efficiency and visualization, may inadvertently damage cells by rupturing membranes or disrupting nuclei, thereby reducing the yield and integrity of cellular material available for analysis. Given the critical importance of maximizing diagnostic sensitivity in a disease with high morbidity and mortality, refining vitrectomy parameters to minimize cellular damage could represent a major advancement in the diagnostic workflow of VRL. The present study aims to systematically evaluate the effect of different combinations of vacuum pressure and cut rate on cellular recovery and morphological preservation in a controlled ex-vivo vitreous model. Approximately 100 healthy donors will be enrolled and stratified into four experimental groups, each undergoing simulated diagnostic vitrectomy procedures under distinct parameter settings: * Group A: Low vacuum / Low cut rate * Group B: Low vacuum / High cut rate * Group C: High vacuum / Low cut rate * Group D: High vacuum / High cut rate This design enables direct comparison of operational conditions to identify settings that optimize cell preservation and recovery. The ultimate goal is to inform best-practice recommendations for diagnostic vitrectomy in suspected VRL, potentially improving cytologic yield and overall diagnostic accuracy.
Study Type
OBSERVATIONAL
Enrollment
100
The simulated vitrectomy is performed using the cutter handpiece, a microcannulated probe that combines a pneumatic cutting mechanism and a controlled aspiration system. The probe is directly inserted into the collection tube (ex-vivo model), allowing mechanical fragmentation and aspiration of donor suspended cells under the selected physical parameters (vacuum and cut rate, according to protocol).
Difference in absolute lymphocyte count between post-processing and baseline measurements (Δ lymphocyte count).
To quantify the impact of different vitrectomy parameter combinations (vacuum pressure and cut rate) on lymphocyte recovery after ex-vivo processing of whole blood using a vitreous trap model.
Time frame: Same-day assessment, immediately after processing
Δ values for the listed CBC parameters
To quantify the pre/post variation (Δ) in complete blood count (CBC) parameters - including WBC, RBC, PLT, HGB, HCT, MCV, MCH, MCHC, NEUT, MONO, EO, and BASO. To compare outcomes across the four parameter combinations in order to identify the settings that best preserve cellular integrity and recovery.
Time frame: Same-day assessment, immediately after processing
Semiquantitative cytology integrity score (smear evaluation)
To assess differences in cytological integrity on optical smears across the four parameter combinations in order to identify the settings that best preserve cellular integrity and recovery.
Time frame: Same-day assessment, immediately after processing
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