The use of collagenase and Dispase does not increase or diversify the population type of macrophages when extracting bone marrow cells from mice
Faculty Mentor
Jason Ashley
Presentation Type
Poster
Start Date
May 2025
End Date
May 2025
Location
PUB NCR
Primary Discipline of Presentation
Biology
Abstract
Osteoclasts, responsible for bone resorption, differentiate from Bone Marrow Hematopoietic Stem Cells (BM-HSCs). Bone flushing is the standard method for isolating bone marrow macrophages, but some studies suggest that bone crushing with collagenase and dispase may yield a higher cell count and diversity. This study investigates whether this enzymatic method enhances macrophage population recovery compared to traditional bone flushing. Bone marrow was extracted using 10 mL of Alpha 10 medium, with 1.25 mL applied to each end of four bones. This yielded adherent BM-MSCs and stromal cells, as well as non-adherent macrophages and red blood cells. Some protocols utilize collagenase and dispase to digest bone in an effort to increase cell yield and diversity. Trypan blue counting assessed cell density, while flow cytometry analyzed cell size and organelle density for population differences. Collagenase and dispase did not significantly increase cell density or alter population diversity. Flow cytometry forward and backward scatter plots showed identical distributions between methods. The enzymatic digestion method does not provide a significant advantage in cell recovery or diversity. Given its additional two-hour processing time, it is inefficient. Bone flushing remains the most effective and time-efficient method for isolating bone marrow cells.
Recommended Citation
Mulch, Sabin Todd; Boruff, Maxwell; and Quinn, Colton, "The use of collagenase and Dispase does not increase or diversify the population type of macrophages when extracting bone marrow cells from mice" (2025). 2025 Symposium. 16.
https://dc.ewu.edu/srcw_2025/ps_2025/p2_2025/16
Creative Commons License
This work is licensed under a Creative Commons Attribution-NonCommercial-No Derivative Works 4.0 International License.
The use of collagenase and Dispase does not increase or diversify the population type of macrophages when extracting bone marrow cells from mice
PUB NCR
Osteoclasts, responsible for bone resorption, differentiate from Bone Marrow Hematopoietic Stem Cells (BM-HSCs). Bone flushing is the standard method for isolating bone marrow macrophages, but some studies suggest that bone crushing with collagenase and dispase may yield a higher cell count and diversity. This study investigates whether this enzymatic method enhances macrophage population recovery compared to traditional bone flushing. Bone marrow was extracted using 10 mL of Alpha 10 medium, with 1.25 mL applied to each end of four bones. This yielded adherent BM-MSCs and stromal cells, as well as non-adherent macrophages and red blood cells. Some protocols utilize collagenase and dispase to digest bone in an effort to increase cell yield and diversity. Trypan blue counting assessed cell density, while flow cytometry analyzed cell size and organelle density for population differences. Collagenase and dispase did not significantly increase cell density or alter population diversity. Flow cytometry forward and backward scatter plots showed identical distributions between methods. The enzymatic digestion method does not provide a significant advantage in cell recovery or diversity. Given its additional two-hour processing time, it is inefficient. Bone flushing remains the most effective and time-efficient method for isolating bone marrow cells.
