Background/Purpose: There are many types of particulate matter in synovial fluid, and specific identification of these particles are challenging. We assessed synovial fluid samples from arthropathic joints and identified that crystals displaying Maltese-cross pattern under polarized light microscopy. We are first to report that such crystals are actually proteinaceous amorphous calcium carbonate crystals (PACC).

Methods: 174 symptomatic synovial fluid (SF) samples were included in this study, which were collected as discarded SF for joints aspirated for clinical care. Polarized light microscopy and alizarin red S staining were used to screen SF samples. Chemically specific techniques including Raman microspectroscopy (μRS) and scanning electron microscopy (SEM) coupled with energy dispersive x-ray (EDX) analyses were used to characterize crystals with Maltese-cross patterned birefringence.

Results: Maltese-cross birefringent crystals were present in 5.7% (10 of 174 SF) of symptomatic joints and positively stained with alizarin red S. Raman spectrum of these crystals was similar to those of sodium carbonate and calcium carbonate in bands 700 – 750 cm^-1 and 1070 – 1090 cm^-1. Raman microspectroscopy (μRS) and SEM/EDX analyses showed absence of phosphate groups, excluding the possible crystals’ association with basic calcium phosphates. The molar ratio (1:3.68) between calcium and oxygen in these crystals was close to the ratio in calcium carbonate (CaCO₃, 1:3). These crystals dissolved with protein digesting enzymes. As a result, we term these crystals “proteinaceous calcium carbonate crystals (PCCD)”.

Conclusion: Theses data provide chemically specific evidence that birefringent particles displaying Maltese-cross birefringence patterns are an amorphous form of calcium carbonate that also include a proteinaceous phase. Our findings counter the prior notion that such crystals are lipids (based on birefringence) or basic calcium phosphates (based on alizarin red S staining). These crystals were observed in symptomatic joints, occurred at a frequency comparable to that observed for calcium pyrophosphate (CPPD) crystals, and their presence was initially overlooked by clinical microscopic observation. This study should elevate the consciousness of clinical microscopists in considering PACC as a crystal species involved in joint arthropathy.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/proteinaceous-amorphous-calcium-carbonates-as-a-novel-family-of-crystals-in-synovial-fluid-from-symptomatic-joints/