library(tidytof)
library(dplyr)
library(ggplot2)
count <- dplyr::count
Often, high-dimensional cytometry experiments collect tens or hundreds or millions of cells in total, and it can be useful to downsample to a smaller, more computationally tractable number of cells - either for a final analysis or while developing code.
To do this, {tidytof} implements the tof_downsample() verb, which allows downsampling using 3 methods: downsampling to an integer number of cells, downsampling to a fixed proportion of the total number of input cells, or downsampling to a fixed cellular density in phenotypic space.
Downsampling with tof_downsample()
Using {tidytof}’s built-in dataset phenograph_data, we can see that the original size of the dataset is 1000 cells per cluster, or 3000 cells in total:
data(phenograph_data)
phenograph_data |>
dplyr::count(phenograph_cluster)
#> # A tibble: 3 × 2
#> phenograph_cluster n
#> <chr> <int>
#> 1 cluster1 1000
#> 2 cluster2 1000
#> 3 cluster3 1000
To randomly sample 200 cells per cluster, we can use tof_downsample() using the “constant” method:
phenograph_data |>
# downsample
tof_downsample(
group_cols = phenograph_cluster,
method = "constant",
num_cells = 200
) |>
# count the number of downsampled cells in each cluster
count(phenograph_cluster)
#> # A tibble: 3 × 2
#> phenograph_cluster n
#> <chr> <int>
#> 1 cluster1 200
#> 2 cluster2 200
#> 3 cluster3 200
Alternatively, if we wanted to sample 50% of the cells in each cluster, we could use the “prop” method:
phenograph_data |>
# downsample
tof_downsample(
group_cols = phenograph_cluster,
method = "prop",
prop_cells = 0.5
) |>
# count the number of downsampled cells in each cluster
count(phenograph_cluster)
#> # A tibble: 3 × 2
#> phenograph_cluster n
#> <chr> <int>
#> 1 cluster1 500
#> 2 cluster2 500
#> 3 cluster3 500
And finally, we might also be interested in taking a slightly different approach to downsampling that reduces the number of cells not to a fixed constant or proportion, but to a fixed density in phenotypic space. For example, the following scatterplot demonstrates that there are certain areas of phenotypic density in phenograph_data that contain more cells than others along the cd34/cd38 axes:
rescale_max <-
function(x, to = c(0, 1), from = range(x, na.rm = TRUE)) {
x / from[2] * to[2]
}
phenograph_data |>
# preprocess all numeric columns in the dataset
tof_preprocess(undo_noise = FALSE) |>
# plot
ggplot(aes(x = cd34, y = cd38)) +
geom_hex() +
coord_fixed(ratio = 0.4) +
scale_x_continuous(limits = c(NA, 1.5)) +
scale_y_continuous(limits = c(NA, 4)) +
scale_fill_viridis_c(
labels = function(x) round(rescale_max(x), 2)
) +
labs(
fill = "relative density"
)
To reduce the number of cells in our dataset until the local density around each cell in our dataset is relatively constant, we can use the “density” method of tof_downsample:
phenograph_data |>
tof_preprocess(undo_noise = FALSE) |>
tof_downsample(method = "density", density_cols = c(cd34, cd38)) |>
# plot
ggplot(aes(x = cd34, y = cd38)) +
geom_hex() +
coord_fixed(ratio = 0.4) +
scale_x_continuous(limits = c(NA, 1.5)) +
scale_y_continuous(limits = c(NA, 4)) +
scale_fill_viridis_c(
labels = function(x) round(rescale_max(x), 2)
) +
labs(
fill = "relative density"
)
Thus, we can see that the density after downsampling is more uniform (though not exactly uniform) across the range of cd34/cd38 values in phenograph_data.
Additional documentation
For more details, check out the documentation for the 3 underlying members of the tof_downsample_* function family (which are wrapped by tof_downsample):
tof_downsample_constanttof_downsample_proptof_downsample_density
Session info
sessionInfo()
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#> Running under: Ubuntu 22.04.4 LTS
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