A Micro-Optic Stalk (μOS) System to Model the Collective Migration of Retinal Neuroblasts

Round 1

Reviewer 1 Report

The manuscript entitled “A micro optic stalk (μOS) system to model the collective migration of retinal neuroblasts“ by Zhang et al. presents a microfluidic device for modeling the collective migration of retinal neuroblasts. The authors presented some interesting results using primary Drosophila cells, yet the presented method is not really new. It is recommended that the authors should better argue the novelty of this work and revise the article. The detailed comments are listed below:

There are many previous microfluidic devices having similar geometry for cell migration and chemo-gradient formation. The presented method is not really new. It is recommended that the authors should better introduce relevant previous works in this field. Then, the authors should argue the novelty of the presented platform. Some related articles are attached:

"Polar stimulation and constrained cell migration in microfluidic channels"
"Single-cell migration chip for chemotaxis-based microfluidic selection of heterogeneous cell populations"
"Cell migration in microengineered tumor environments"
"Microfluidic Collective Cell Migration Assay for Study of Endothelial Cell Proliferation and Migration under Combinations of Oxygen Gradients, Tensions, and Drug Treatments"

It is recommended that the authors can better explain the culture environment (e.g. temperature, media, and CO2 level) of Primary RNBs. Will that affect the migration results?

It is recommended that the authors can add a schematic to explain where the cells were loaded and their migratory direction. It would be easier for the audience to follow.

The migration distance presented in Table 2 is pretty short. Is that the cell movement after 6-8 hours? It is recommended that the authors can better explain those data.

Following the previous question, it is recommended the authors can monitor the movement of a cell/cluster to verify its movement process.

From the Fig. 6A, it seems like the cell adhesion on the substrate is not great. That can significantly affect the cell migration speed. It is recommended that the authors can test different coating conditions on the substrate. 

Author Response

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Author Response File: Author Response.pdf

Reviewer 2 Report

This manuscript reports on the use of a microfluidic device for investigating the collective migration behavior of retinal neuroblasts (RNBs). The device concept is similar to that used to form synapses. The results suggest that small clusters migrate faster than larger clusters and single RNBs are not responsive to the fibroblast growth factor (FGF) gradient. Overall, the idea of the manuscript is interesting. However, it needs significant improvement before it can be considered for publication.

 

1. Results 3.1 the design shall part of the methods.
2. The dimensions of the device are not clearly marked. In fact, it is very confusing. The authors are recommended to provide a cross-sectional view schematic of the Y channel from the left or right side. Line 246, the use of “diameters” is very confusing as these channels are essentially rectangular (37 μm × 10 μm or 35 μm ×10 μm) rather than circular in cross-section. If the diameter of the optical stalk is about 35 um, why the channel height was designed to be 10 μm? What are the “two reservoirs”? Are these the side channels on the left and right sides of the OS channels? If so, the spacing of 100 μm does not seem right. Please check and clarify.
3. Line 161, QR and QL shall be defined first.
4. Line 362 What was the flow rate used?
5. Line 379 shall be 3-4 μm instead of 3-4 mm?
6. Line 386 the unit is missing.
7. 3A labels are not clear.
8. 5 E and F are difficult to see the details.
9. What was the migration duration in Table 2?
10. Large clusters disaggregated during observation. Authors observed at 2, 4 and 6 hrs. What was the percentage of small clusters after 6 hrs? Say, 8, 10, 12 hrs? Also, did the authors repeat these experiments? Fig. 6d is very confusing as there is no horizontal axis title. The percentages should have error bars of at least three replicates.
11. 6A-C do not show the disaggregation of large cluster. Authors shall provide images of the initial intact cluster and the disaggregated clusters after.
12. Authors are also recommended to provide time-lapsed images of the motile clusters and those immotile single cells.
13. Check typos, e.g., Line 61 should be “model of the developing”.

Author Response

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Author Response File: Author Response.pdf

Round 2

Reviewer 1 Report

The authors addressed the comments from reviewer.

Author Response

The authors thank the reviewer for their time and suggestions during the submission process.

Reviewer 2 Report

The authors have improved the quality of the manuscript. However, two concerns have not been addressed. 

1. Since the diameter of the optical stalk is about 35 μm, why the channel height was designed to be 10 μm?
2. Why the error bars in the new Fig.6G seem identical? Are these error bars from three replicates?

Author Response

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Author Response File: Author Response.docx