TABLE 1.

Some examples of studies providing single-cell insight into HIV-1 biology or pathogenesis.

	Approaches	Description	Few examples of application
Detection of rare events	Branched DNA signal amplification (RNA or DNA)	Flow cytometric or microscopic detection of RNAs or DNAs, compatible with protein co-detection	Compare latency reversal in different cell subsets (Baxter et al., 2016; Grau-Exposito et al., 2019)
			Quantify and phenotype the viral reservoirs ex vivo (Baxter et al., 2016; Grau-Exposito et al., 2017)
			Interrogate viral reservoirs in tissues (Deleage et al., 2016) and estimate whole body viral burden (Estes et al., 2017)
			Identify HIV+ cells in tissue-resident cells, including non-T cells (Vasquez et al., 2018)
	Dual protein detection	Co-detection of viral proteins by flow cytometry	Study translation-competent viral reservoirs (DeMaster et al., 2015; Pardons et al., 2019)
Genetic profiling	Targeted PCR for viral genes	Quantification of RNA or DNA targets	Correlate residual HIV-1 transcription to sites of integrated proviruses (Wiegand et al., 2017)
			Quantify HIV-1 splicing upon latency reversal (Yucha et al., 2017)
			Assess gene expression in different stages of SIV replication (Bolton et al., 2017)
	Unsupervised sequencing (RNAseq, DNAseq, and ATAC-seq)	Unbiased assessment of transcriptional and epigenetic landscapes	Identify biomarkers of HIV-1 permissiveness (Rato et al., 2017)
			Define quiescent HIV-1 infected cells (Bradley et al., 2018; Golumbeanu et al., 2018), B cell profile post-vaccination (de Armas et al., 2019)
			Establish an epigenetic signature of resident memory T cells during HIV infection (Buggert et al., 2018)
	BCR and TCR sequencing	Profiling of the B cell and T cell repertoires	Analysis of BCR repertoire post-immunization (Scheid et al., 2009; Sundling et al., 2014)
			Study T cell clonal expansion in vivo in the context of HIV infection (Wendel et al., 2018)
	Integration sequencing	Mapping of integrated vDNA	Map HIV-1 integration sites in the CD4+ T cell genome of primary samples (Cohn et al., 2015)
	Virus barcoding	Engineered viruses with degenerate unique barcodes	Examine the transcriptional potential of integrations sites by correlating barcodes in integrated DNA and vRNA (Chen et al., 2017)
High dimensional phenotyping	Mass cytometry (CyTOF)	Time-of-flight cytometry based on heavy ion metal tags with minimal spectral overlap	Evaluate the susceptibility of CD4+ T subsets to productive HIV-1 infection (Cavrois et al., 2017)
			Define the phenotypic landscape of exhausted T cells (Bengsch et al., 2018a; Bekele et al., 2019)
			Link new CD8+ T cell subsets to HIV-1 pathogenesis (Buggert et al., 2018)
Imaging of subcellular molecular dynamics	Fluorescent tags	Temporal interrogation of bioengineered fluorescently tagged proteins of interest in primary cells	Dissect, in live cells viral entry (Miyauchi et al., 2009), uncoating (Arhel et al., 2006; Mamede et al., 2017; Francis and Melikyan, 2018b), nuclear import (Chin et al., 2015), and assembly (Ivanchenko et al., 2009) Estimate the timeline of gene expression (Holmes et al., 2015)
	Branched DNA signal amplification for RNA/DNA single-cell microscopy	Snapshots of selected RNAs, vDNA and proteins sub-localization	Study the nuclear import of vDNA (Chin et al., 2015)
			Locate integration sites of native proviruses in primary cells (Marini et al., 2015)
			Study the uncoating of native viruses (Puray-Chavez et al., 2017)
	Imaging of integrated DNA	SCIP	Investigate the spatial localization of HIV-1 integration sites in live cells (Di Primio et al., 2013)
		Detection of CRISPR-Cas9-cleaved integrated provirus	Assess HIV-1 integration in real-time in live cells (Ma et al., 2017)