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  • Clinical Research Article
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Umbilical cord milking as a technique to harvest cord blood derived cells for regenerative applications

Pediatric Research (2025)Cite this article

Abstract

Background

Umbilical cord blood (UCB) is increasingly studied for regenerative therapies, yet the impact of different collection techniques on cell yield and quality remains unclear. This study compared standard needle-and-bag UCB collection with manual cord milking, performed both in utero (placenta attached) and ex utero (placenta delivered), using samples from healthy term infants ( > 37 weeks gestation).

Method

Forty-two samples (n = 10 standard in utero, n = 10 standard ex utero, n = 10 milking in utero, n = 12 milking ex utero) were analyzed for blood volume, mononuclear cell count, and cellular composition via flow cytometry. Key cell populations included hematopoietic stem cells (CD34 + CD45 + ), endothelial progenitor cells (CD45 + CD34 + CD31 − CD133 + ), and mature endothelial cells (CD34 − CD45 − CD31 + ). Plasma cytokines, including inflammatory and angiogenic markers, were also assessed.

Results

No significant differences were found in total blood volume or mononuclear cell counts across groups. However, endothelial progenitor cell viability was significantly reduced in cord milking ex utero compared to standard in utero collection (p < 0.0001). Cytokine analysis showed elevated IL-1RA and reduced VEGF-A in cord milking ex utero samples (p < 0.0001 and p = 0.0004, respectively).

Conclusion

These findings suggest that in utero cord milking may be a viable alternative to standard UCB collection, preserving cell viability and cytokine integrity.

Impact

  • The standard method for collecting umbilical cord blood (UCB) has limitations, especially in cases like premature birth, where low volumes yield insufficient mononuclear and hematopoietic stem cells for therapeutic use.

  • This study evaluated an alternative technique—umbilical cord milking—against the standard approach.

  • As the first study to assess its efficacy for UCB collection, the findings offer insights into a viable alternative method.

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Fig. 1: UCB cell phenotype analysis.
Fig. 2: UCB cell viability obtained through flow cytometry analysis.
Fig. 3: Inflammatory and angiogenic marker analysis of UCB.

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Data availability

All data is available on reasonable request from authors.

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Funding

No specific funding was secured for this research. R.A.S.M. and A.R. are supported by PhD Scholarships from Monash University. A.R. receives funding from Lions Cord Blood Foundation and Royal Australasian College of Physicians. A.M. and C.A.M. receive funding for their research from the National Health and Medical Research Council of Australia. T.R.P., A.M., and C.A.M. receive funding from the Medical Research Future Fund. A.M. also receives funding from Monash Health Foundation, National Stem Cell Foundation of Australia, Lions Cord Blood Foundation and Cerebral Palsy Alliance.

Author information

Author notes

  1. These authors contributed equally: Courtney A. McDonald, Atul Malhotra.

Authors and Affiliations

  1. The Ritchie Centre, Hudson Institute of Medical Research, Melbourne, VIC, Australia

    Ricki Anne S. Marzan, Abdul Razak, Lindsay Zhou, Tayla R. Penny, Courtney A. McDonald & Atul Malhotra

  2. Department of Obstetrics and Gynecology, Monash University, Melbourne, VIC, Australia

    Ricki Anne S. Marzan, Tayla R. Penny & Courtney A. McDonald

  3. Department of Paediatrics, Monash University, Melbourne, VIC, Australia

    Kristyn Connelly, Abdul Razak, Lindsay Zhou & Atul Malhotra

  4. Monash Newborn, Monash Children’s Hospital, Melbourne, VIC, Australia

    Kristyn Connelly, Abdul Razak, Lindsay Zhou & Atul Malhotra

Authors
  1. Ricki Anne S. Marzan
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  2. Kristyn Connelly
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  4. Lindsay Zhou
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Contributions

R.A.S.M.: Conduct of experiments, data collection and analysis, interpretation, manuscript writing. K.C.: recruitment, conduct of experiments, data collection, manuscript editing. A.R.: recruitment, conduct of experiments, data collection and analysis, manuscript editing. L.Z.: recruitment, conduct of experiments, data collection, manuscript editing. T.R.P.: conception and design, conduct of experiments, data analysis, manuscript editing, supervision. C.A.M.: conception and design, conduct of experiments, data analysis and interpretation, manuscript editing, supervision. A.M.: conception and design, data analysis and interpretation, manuscript editing, supervision.

Corresponding author

Correspondence to Atul Malhotra.

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Marzan, R.A.S., Connelly, K., Razak, A. et al. Umbilical cord milking as a technique to harvest cord blood derived cells for regenerative applications. Pediatr Res (2025). https://doi.org/10.1038/s41390-025-04687-6

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