Page 193 - IJB-10-2

International Journal of Bioprinting Optimizing inkjet bioprinting

and varies by cell type. The model was further extended 2.2. Cell homogeneity within bio-inks
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to describe viscosity in concentrated solutions with the Cell sedimentation is a common occurrence in most inkjet
following equation: bioprinting systems, primarily caused by the influence
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of gravitational forces on the freely suspended cells.
25/ 25 . The uniformity of cell distribution within the printing
1 0 s 0 (III)
25 . cartridge changes with time as these suspended cells
s
settle to the bottom of the bio-ink reservoir, resulting
This model is only valid where cell membrane surface in cell inhomogeneity in the printing output (Figure
tension is sufficient to keep the cell mostly spherical. A 2b). A research study has investigated the impact of
different model for viscosity of a concentrated suspension cell sedimentation on the bioprinter’s output and has
of poly-dispersed spheres has been proposed. highlighted the issue of inconsistent printing output over
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time. As time progresses, the concentration of localized
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25/ cells at the bottom of the reservoir increases significantly,
1 0 s (IV) which can lead to problems such as nozzle clogging and
0 highly unstable droplet ejection. Consequently, this results
s
Notably, this model does not explicitly account for size in a decline in overall printing reliability and quality.
polydispersity. For high-volume fraction cell suspensions, a The cell homogeneity within the bio-inks can be
more comprehensive model of model treats the suspension examined by recording real-time video of the sedimentation
as a viscoelastic fluid. The viscous response is attributed process and calculating the cell sedimentation velocity
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to the cytoplasmic viscosity and the viscous contribution of using sophisticated optical set-ups. The theoretical values
the cell membrane, while the elastic response is attributed for cell sedimentation velocity can be determined using
to the cell membrane’s ability to reversibly restore the cell Stokes’ law, defined as follows:
to a spherical morphology. In situations where cells at high
volume fractions tend to aggregate, these aggregates also ( ) gD 2
exhibit viscoelastic behavior, contributing to the overall v p f C (VI)
viscoelastic response of the suspension. This behavior can 18
be modeled by the semi-empirical Mooney equation:
where v represents the sedimentation velocity, g is the
gravitational acceleration, D is the cell diameter, ρ and
c
p

K ρ are the densities of particles and the fluid, respectively,
f
0 exp 1 (V) and µ is the fluid viscosity. As the distance between
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1 K neighboring cells decreases, the cells start adhering to each
2 D other and form larger cell aggregates, which can exceed
In this equation, K and K are fitting parameters that the nozzle diameter and result in nozzle clogs and unstable
1
2
consider cell shape and the degree of cell aggregation; γ ⋅ droplet ejection. Additionally, interactions such as van der
represents the fluid shear rate, b is the shear flow index; σ is Waals forces between the inner surfaces of the printing
the effective cell membrane thickness, and D is the average cartridge and the suspended cells can cause cells to adhere
cell diameter. Typically, K , K , and b are determined to the walls; this is especially important in constrictions
2
1
experimentally by measuring the complex viscosity as a in the print cartridge, where the adhered cells can grow
function of shear rate in a rheometer. into larger cell aggregates and clog the constriction. 36,37
The presence of particles (including cells) in high These floating cells are then attracted to the adhered cells,
concentration (>0.5% v/v) decreases surface tension, and contributing to the growth of larger cell aggregates near the
surface tension decreases asymptotically with increasing constriction at the bottom of the printing cartridge.
particle loading. However, in practical bio-inks, other To counter cell sedimentation and minimize cell
substances including peptides, proteins, and cellular adhesion and aggregation along the cartridge’s constricted
debris have a substantially dominating effect of lowering regions, an inert polyvinylpyrrolidone (PVP)-based bio-
the surface tension. Thus, in practice, surface tension of ink can be used to pre-coat the printing cartridge. Other
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bio-inks is determined empirically. By understanding strategies to mitigate cell sedimentation and aggregation
the influence on cells on the physical properties of the include active stirring and modifying the bio-ink
bio-inks, one can formulate a cell-laden bio-ink of high properties to achieve neutral buoyancy. It is important to
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printability. note that while active stirring can be effective in reducing

Volume 10 Issue 2 (2024) 185 doi: 10.36922/ijb.2135