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Purpose <p>Cultivation of <i>Theileria annulata</i> schizont-transformed cell lines typically relies on fetal bovine serum (FBS), limiting standardization. This study evaluated three serum-free media (SFMs) for the in-vitro cultivation of <i>T. annulata</i>-transformed cell lines derived from different isolates.</p> Methods <p>TaNM1 and TaXJS cell lines were adapted to AIM-V, VP-SFM, and HL-1, and compared against FBS-supplemented RPMI-1640. Proliferation, apoptosis, and parasite burden were assessed via viable cell counting, flow cytometry, and a novel <i>TaSP</i>-targeted absolute quantitative PCR (qPCR).</p> Results <p>AIM-V SFM and VP-SFM supported sustained proliferation of both cell lines after adaptation. AIM-V SFM showed the most stable performance among the tested serum-free conditions, with mean doubling times of 35.45&#xa0;h for TaNM1 and 52.45&#xa0;h for TaXJS. In contrast, unsupplemented HL-1 did not support robust growth, whereas supplementation with hypoxanthine-thymidine(HT) and AlbuMax I partially restored growth. Flow cytometry showed no marked increase in apoptosis under serum-free conditions, and qPCR indicated maintenance of <i>T. annulata</i> genome copy numbers during serum-free culture.</p> Conclusions <p>Serum-free culture of <i>T. annulata</i> schizont-transformed cell lines is feasible but strongly medium-dependent. AIM-V SFM and VP-SFM supported serum-free maintenance after adaptation, whereas HL-1 required supplementation. These findings provide a basis for further optimization of serum-free culture systems for standardized research and vaccine-related applications.</p>

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Optimization of Serum-Free Culture Medium for Theileria annulata-Transformed Cell Lines: A Comparative Analysis

  • 期刊论文

  • 发表日期：

    2026年5月11日

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  • Peiqi Liu,
  • Shuaiyang Zhao,
  • Jiali Yao,
  • Zeen Liu,
  • Muhammad Kashif Obaid,
  • Hong Yin,
  • Guiquan Guan

摘要

Purpose

Cultivation of Theileria annulata schizont-transformed cell lines typically relies on fetal bovine serum (FBS), limiting standardization. This study evaluated three serum-free media (SFMs) for the in-vitro cultivation of T. annulata-transformed cell lines derived from different isolates.

Methods

TaNM1 and TaXJS cell lines were adapted to AIM-V, VP-SFM, and HL-1, and compared against FBS-supplemented RPMI-1640. Proliferation, apoptosis, and parasite burden were assessed via viable cell counting, flow cytometry, and a novel TaSP-targeted absolute quantitative PCR (qPCR).

Results

AIM-V SFM and VP-SFM supported sustained proliferation of both cell lines after adaptation. AIM-V SFM showed the most stable performance among the tested serum-free conditions, with mean doubling times of 35.45 h for TaNM1 and 52.45 h for TaXJS. In contrast, unsupplemented HL-1 did not support robust growth, whereas supplementation with hypoxanthine-thymidine(HT) and AlbuMax I partially restored growth. Flow cytometry showed no marked increase in apoptosis under serum-free conditions, and qPCR indicated maintenance of T. annulata genome copy numbers during serum-free culture.

Conclusions

Serum-free culture of T. annulata schizont-transformed cell lines is feasible but strongly medium-dependent. AIM-V SFM and VP-SFM supported serum-free maintenance after adaptation, whereas HL-1 required supplementation. These findings provide a basis for further optimization of serum-free culture systems for standardized research and vaccine-related applications.