IFNc/IL-10 Co-producing Cells Dominate the CD4
Response to Malaria in Highly Exposed Children
Prasanna Jagannathan1, Ijeoma Eccles-James1, Katherine Bowen1, Felistas Nankya2, Ann Auma2,
Samuel Wamala2, Charles Ebusu2, Mary K. Muhindo2, Emmanuel Arinaitwe2, Jessica Briggs1,
Bryan Greenhouse1, Jordan W. Tappero3, Moses R. Kamya4, Grant Dorsey1, Margaret E. Feeney1,5*
1Department of Medicine, San Francisco General Hospital, University of California, San Francisco, San Francisco, California, United States of America, 2Infectious Diseases
Research Collaboration, Kampala, Uganda, 3Center for Global Health, Centers for Disease Control and Prevention, Atlanta, Georgia, United States of America,
4Department of Medicine, Makerere University College of Health Sciences, Kampala, Uganda, 5Department of Pediatrics, University of California, San Francisco, San
Francisco, California, United States of America
Although evidence suggests that T cells are critical for immunity to malaria, reliable T cell correlates of exposure to and
protection from malaria among children living in endemic areas are lacking. We used multiparameter flow cytometry to
perform a detailed functional characterization of malaria-specific T cells in 78 four-year-old children enrolled in a
longitudinal cohort study in Tororo, Uganda, a highly malaria-endemic region. More than 1800 episodes of malaria were
observed in this cohort, with no cases of severe malaria. We quantified production of IFNc, TNFa, and IL-10 (alone or in
combination) by malaria-specific T cells, and analyzed the relationship of this response to past and future malaria incidence.
CD4+T cell responses were measurable in nearly all children, with the majority of children having CD4+T cells producing
both IFNc and IL-10 in response to malaria-infected red blood cells. Frequencies of IFNc/IL10 co-producing CD4+T cells,
which express the Th1 transcription factor T-bet, were significantly higher in children with $2 prior episodes/year compared
to children with ,2 episodes/year (P,0.001) and inversely correlated with duration since malaria (Rho=20.39, P,0.001).
Notably, frequencies of IFNc/IL10 co-producing cells were not associated with protection from future malaria after
controlling for prior malaria incidence. In contrast, children with ,2 prior episodes/year were significantly more likely to
exhibit antigen-specific production of TNFa without IL-10 (P=0.003). While TNFa-producing CD4+T cells were not
independently associated with future protection, the absence of cells producing this inflammatory cytokine was associated
with the phenotype of asymptomatic infection. Together these data indicate that the functional phenotype of the malaria-
specific T cell response is heavily influenced by malaria exposure intensity, with IFNc/IL10 co-producing CD4+T cells
dominating this response among highly exposed children. These CD4+T cells may play important modulatory roles in the
development of antimalarial immunity.
Citation: Jagannathan P, Eccles-James I, Bowen K, Nankya F, Auma A, et al. (2014) IFNc/IL-10 Co-producing Cells Dominate the CD4 Response to Malaria in Highly
Exposed Children. PLoS Pathog 10(1): e1003864. doi:10.1371/journal.ppat.1003864
Editor: Jean Langhorne, National Institute for Medical Research, United Kingdom
Received August 22, 2013; Accepted November 19, 2013; Published January 9, 2014
This is an open-access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for
any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication.
Funding: This work was supported by the Centers for Disease Control and Prevention (Cooperative Agreement No U62P024421); NIH/NIAID R01AI093615 (MEF),
UCSF Centers for AIDS Research (Supplement to MEF, P30AI027763), NIH/NIAID U19AI089674 (GD), NIH/NIAID K23 AI100949 (PJ), and Burroughs Wellcome Fund/
American Society of Tropical Medicine and Hygiene (PJ). Additional support was provided by the National Center for Advancing Translational Sciences/NIH,
through UCSF-CTSI Grant Number UL1 TR000004. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the
manuscript. The findings and conclusions in this paper are those of the authors and do not necessarily represent the official position of the Centers for Disease
Control and Prevention or the NIH.
Competing Interests: The authors have declared that no competing interests exist.
* E-mail: email@example.com
Clinical immunity to malaria eventually develops in endemic
populations, but only after repeated infections with significant
morbidity to both individuals and their communities . Studies in
regions of high malaria transmission intensity have consistently
shown that the incidence of severe disease decreases considerably
after the first years of life, but sterile immunity (i.e. protection
against parasitemia) develops rarely if ever [2,3]. Moreover,
previously immune individuals may lose protection against symp-
tomatic infection in the absence of continuous exposure [4,5]. The
reasons underlying the slow acquisition of clinical immunity and the
failure to develop sterilizing immunity are unclear, but may include
responses [7–12], and/or host immunoregulatory mechanisms
induced by the parasite [13–19]. As the incidence of malaria
continues to be high in many parts of Africa despite insecticide-
treated bednets and artemisinin-based combination therapy [20–
22], there is a tremendous need to better understand mechanisms of
immunity to malaria in naturally exposed populations. The
identification of immunologic correlates of exposure and protection
in naturally exposed children would significantly help with the
rational design of vaccines and other malaria control interventions.
Both CD4+and CD8+T cells have been demonstrated to play
an important role in protective antimalarial immunity in mouse
models [23–30], and experimental challenge models in humans
and mice strongly suggest that malaria-specific T cells contribute
to protective immunity [31–36]. However, the identification of T
cell correlates of immunity in field-based studies of naturally
exposed humans has proven to be quite challenging. Prior studies
PLOS Pathogens | www.plospathogens.org1 January 2014 | Volume 10 | Issue 1 | e1003864
employing cross-sectional or prospective cohort designs have
found associations between cellular immune responses and
protection from future malaria, including IFNc responses to liver
stage [37–40] and/or merozoite stage malaria antigens [41–44].
However, such studies may be confounded by the level of exposure
to malaria-infected mosquitoes, which varies greatly within
populations, leading subjects with lower exposure to be mis-
categorized as ‘‘protected’’ [45,46]. Because naturally acquired
immunity confers relative rather than absolute protection –
manifested by a gradual decline in the incidence of clinical disease
- careful quantitative outcome measures are essential, but few
population-based studies of natural immunity have included
careful measurement of malaria incidence over time.
Pathogen-specific T cells exhibit notable functional heteroge-
neity, largely dependent on the antigen and cytokine microenvi-
ronment encountered during activation, and measurement of a
single parameter of T cell function (i.e. IFNc production) may
overlook others that are more critical for protection . In other
parasitic infections such as leishmania [48,49] and toxoplasma
, the functional phenotype of the CD4+T cell response
correlates with the success or failure to clear the pathogen. Recent
observations in individuals naturally exposed to malaria suggest an
important role for CD4+T cell production of TNFa, with or
without IFNc, as a potential immunologic correlate of protection
. Conversely, CD4+T cell production of the regulatory
cytokine IL-10 has been implicated in modulating the severity of
disease [18,52] and may interfere with the development of
protective immunity [14,42,53]. The role of these inflammatory
and regulatory cytokines in mediating protective immunity in
naturally exposed children, and in determining the balance
between immunopathology and chronic repeated infection,
In this study we performed a detailed functional characteriza-
tion of malaria-specific T cell responses among four-year-old
children residing in a highly malaria-endemic region to determine
whether naturally acquired T cell responses correlate with
exposure to and/or protection from malaria. We hypothesized
that CD4+T cells producing the pro-inflammatory cytokines IFNc
and/or TNFa are associated with protection from malaria, and
that T cell production of the regulatory cytokine IL-10 may
interfere with the acquisition of protection. Our results suggest that
the functional phenotype of the malaria-specific T cell response
was heavily influenced by prior malaria exposure intensity, with
CD4+T cells co-producing IFNc and IL10 dominating this
response among highly exposed children. However, these IFNc/
IL-10 co-producing cells were not independently associated with
protection from future malaria, and may be associated with
Study population and clinical outcomes
The study cohort consisted of 78 HIV-uninfected children
followed from infancy through 5 years of age (Table 1). Blood for
this study was drawn at four years of age (range 49–51 months),
and 92% of children continued to be followed through 5 years of
age. A total of 1855 incident cases of malaria were observed in this
cohort through 5 years of age. All children were treated promptly
with artemisinin-based combination therapy, and despite the
strikingly high numbers of malaria episodes, only 4 cases of
malaria were deemed ‘‘complicated’’ (all based on a single
convulsion). No cases of severe malaria (including severe anemia)
were observed. Among children with a lower prior incidence of
malaria (,2 episodes per person year (ppy) between 1 and 4 years
of age, n=10), 90% lived in town; whereas among children with
higher prior malaria incidence (.=2 episodes ppy, n=68), only
7% of children lived in town. This suggests that children with the
lowest prior incidence had less exposure to malaria-infected
mosquitoes. Episodes of asymptomatic parasitemia were rare in
this cohort (median 1 episode per subject over the entire study
period, IQR 0–4, Table 1) and the incidence of malaria declined
only slightly in the year following the blood draw (from 5.7 to 5.1
episodes ppy), suggesting that effective clinical immunity had not
yet emerged in most children. One child had symptomatic malaria
(parasitemia with a fever requiring treatment) at the time of the
blood draw, and 17 (22%) had blood smears demonstrating
The functional phenotype of malaria-specific CD4+T cells
is influenced by prior malaria incidence
To define the frequency and function of malaria-specific T cell
responses, PBMC were stimulated with malaria-infected red blood
cells (iRBC) and analyzed by flow cytometry for production of
IFNc, IL-10, and TNFa (Fig. 1a). The median frequency of
malaria-specific CD4+T cell responses producing any of these
cytokines, alone or in combination, was 0.20% (IQR 0.12%–
0.35%). Among all children, frequencies of CD4+T cells
producing IFNc (median 0.16%) and IL-10 (median 0.14%) were
significantly higher than those producing TNFa (median 0.04%,
P,0.001, Fig. 1b). Production of these two cytokines largely
overlapped, with a median of 83% of IL-10-producing cells also
making IFNc, and a median of 71% of IFNc-producing cells also
making IL-10. Malaria-specific production of IL-2 was tested in a
subset of children (n=44), but responses were consistently of low
magnitude (median frequency 0.02%, data not shown). At the time
of the assay 17 of the 78 children had positive blood smears;
however there was no significant difference in the overall
frequency of malaria-specific IFNc+(P=0.20), TNFa+(P=0.29),
or IL-10+(P=0.21) CD4+T cells between children with or
without parasitemia. Malaria-specific CD8 T cell responses were
Despite reports of decreasing malaria morbidity across
many parts of Africa, the incidence of malaria among
children continues to be very high in Uganda, even in the
setting of insecticide-treated bednets and artemisinin-
based combination therapy. Additional control measures,
including a vaccine, are sorely needed in these settings,
but progress has been limited by our lack of understand-
ing of immunologic correlates of exposure and protection.
T cell responses to malaria are thought to be important for
protection in experimental models, but their role in
protecting against naturally acquired infection is not clear.
In this study, we performed detailed assessments of the
malaria-specific T cell response among 4-year-old children
living in Tororo, Uganda, an area of high malaria
transmission. We found that recent malaria infection
induces a malaria-specific immune response dominated
by Th1 T cells co-producing IFNc and IL-10, and that these
cells are not associated with protection from future
infection. IFNc/IL-10 co-producing cells have been de-
scribed in several parasitic infections and are hypothesized
to be important in limiting CD4-mediated pathology, but
they may also prevent the development of sterilizing
immunity. These observations have important implications
for understanding the pathophysiology of malaria in
humans and for malaria vaccine development.
CD4 Response to Malaria in Highly Exposed Children
PLOS Pathogens | www.plospathogens.org2January 2014 | Volume 10 | Issue 1 | e1003864
protein correlates with protection from natural Plasmodium falciparum infection
and disease. Nat Med 10: 406–410.
38. Todryk SM, Bejon P, Mwangi T, Plebanski M, Urban B, et al. (2008)
Correlation of memory T cell responses against TRAP with protection from
clinical malaria, and CD4 CD25 high T cells with susceptibility in Kenyans.
PLoS ONE 3: e2027.
39. Kurtis JD, Hollingdale MR, Luty AJ, Lanar DE, Krzych U, et al. (2001) Pre-
erythrocytic immunity to Plasmodium falciparum: the case for an LSA-1
vaccine. Trends Parasitol 17: 219–223.
40. Hoffman SL, Oster CN, Mason C, Beier JC, Sherwood JA, et al. (1989) Human
lymphocyte proliferative response to a sporozoite T cell epitope correlates with
resistance to falciparum malaria. J Immunol 142: 1299–1303.
41. Luty AJ, Lell B, Schmidt-Ott R, Lehman LG, Luckner D, et al. (1999)
Interferon-gamma responses are associated with resistance to reinfection with
Plasmodium falciparum in young African children. J Infect Dis 179: 980–988.
42. Moormann AM, Sumba PO, Chelimo K, Fang H, Tisch DJ, et al. (2013)
Humoral and Cellular Immunity to Plasmodium falciparum Merozoite Surface
Protein 1 and Protection From Infection With Blood-Stage Parasites. J Infect Dis
43. D’Ombrain MC, Robinson LJ, Stanisic DI, Taraika J, Bernard N, et al. (2008)
Association of early interferon-gamma production with immunity to clinical
malaria: a longitudinal study among Papua New Guinean children. Clinical
infectious diseases : an official publication of the Infectious Diseases Society of
America 47: 1380–1387.
44. McCall MB, Hopman J, Daou M, Maiga B, Dara V, et al. (2010) Early
interferon-gamma response against Plasmodium falciparum correlates with
interethnic differences in susceptibility to parasitemia between sympatric Fulani
and Dogon in Mali. J Infect Dis 201: 142–152.
45. Bejon P, Warimwe G, Mackintosh CL, Mackinnon MJ, Kinyanjui SM, et al.
(2009) Analysis of immunity to febrile malaria in children that distinguishes
immunity from lack of exposure. Infect Immun 77: 1917–1923.
46. Greenhouse B, Ho B, Hubbard A, Njama-Meya D, Narum DL, et al. (2011)
Antibodies to Plasmodium falciparum antigens predict a higher risk of malaria
but protection from symptoms once parasitemic. J Infect Dis 204: 19–26.
47. Douek DC, Roederer M, Koup RA (2009) Emerging concepts in the
immunopathogenesis of AIDS. Annual Review of Medicine 60: 471–484.
48. Darrah PA, Patel DT, De Luca PM, Lindsay RW, Davey DF, et al. (2007)
Multifunctional TH1 cells define a correlate of vaccine-mediated protection
against Leishmania major. Nature medicine 13: 843–850.
49. AndersonCF, Oukka M,
CD4(+)CD25(2)Foxp3(2) Th1 cells are the source of IL-10-mediated immune
suppression in chronic cutaneous leishmaniasis. J Exp Med 204: 285–297.
50. Jankovic D, Kullberg MC, Feng CG, Goldszmid RS, Collazo CM, et al. (2007)
Conventional T-bet(+)Foxp3(2) Th1 cells are the major source of host-
protective regulatory IL-10 during intracellular protozoan infection. J Exp
Med 204: 273–283.
51. Olotu A, Moris P, Mwacharo J, Vekemans J, Kimani D, et al. (2011)
Circumsporozoite-Specific T Cell Responses in Children Vaccinated with
RTS,S/AS01(E) and Protection against P falciparum Clinical Malaria. PLoS
ONE 6: e25786.
52. Walther M, Jeffries D, Finney OC, Njie M, Ebonyi A, et al. (2009) Distinct roles
for FOXP3 and FOXP3 CD4 T cells in regulating cellular immunity to
uncomplicated and severe Plasmodium falciparum malaria. PLoS Pathog 5:
53. Zhang G, Manaca MN, McNamara-Smith M, Mayor A, Nhabomba A, et al.
(2012) Interleukin-10 (IL-10) polymorphisms are associated with IL-10
production and clinical malaria in young children. Infect Immun 80: 2316–
54. Epstein JE, Tewari K, Lyke KE, Sim BK, Billingsley PF, et al. (2011) Live
Attenuated Malaria Vaccine Designed to Protect through Hepatic CD8+ T Cell
55. Olotu A, Moris P, Mwacharo J, Vekemans J, Kimani D, et al. (2011)
Circumsporozoite-specific T cell responses in children vaccinated with RTS,S/
AS01E and protection against P falciparum clinical malaria. PLoS ONE 6:
56. Cope A, Le Friec G, Cardone J, Kemper C (2011) The Th1 life cycle: molecular
control of IFN-gamma to IL-10 switching. Trends in Immunology 32: 278–286.
57. O’Garra A, Vieira P (2007) T(H)1 cells control themselves by producing
interleukin-10. Nature reviews Immunology 7: 425–428.
58. Wenisch C, Parschalk B, Narzt E, Looareesuwan S, Graninger W (1995)
Elevated serum levels of IL-10 and IFN-gamma in patients with acute
Plasmodium falciparum malaria. Clinical Immunology and Immunopathology
59. Peyron F, Burdin N, Ringwald P, Vuillez JP, Rousset F, et al. (1994) High levels
of circulating IL-10 in human malaria. Clinical and experimental immunology
60. Wilson NO, Bythwood T, Solomon W, Jolly P, Yatich N, et al. (2010) Elevated
levels of IL-10 and G-CSF associated with asymptomatic malaria in pregnant
women. Infectious Diseases in Obstetrics and Gynecology 2010: pii: 317430.
61. Pinzon-Charry A, Woodberry T, Kienzle V, McPhun V, Minigo G, et al. (2013)
Apoptosis and dysfunction of blood dendritic cells in patients with falciparum
and vivax malaria. J Exp Med 210: 1635–1646.
KuchrooVJ, SacksD (2007)
62. O’Connor RA, Jenson JS, Osborne J, Devaney E (2003) An enduring
association? Microfilariae and immunosuppression [correction of immunosu-
pression] in lymphatic filariasis. Trends Parasitol 19: 565–570.
63. Steel C, Guinea A, McCarthy JS, Ottesen EA (1994) Long-term effect of
prenatal exposure to maternal microfilaraemia on immune responsiveness to
filarial parasite antigens. Lancet 343: 890–893.
64. Wammes LJ, Hamid F, Wiria AE, Wibowo H, Sartono E, et al. (2012)
Regulatory T cells in human lymphatic filariasis: stronger functional activity in
microfilaremics. PLoS Neglected Tropical Diseases 6: e1655.
65. Maizels RM, Yazdanbakhsh M (2003) Immune regulation by helminth parasites:
cellular and molecular mechanisms. Nature reviews Immunology 3: 733–744.
66. McNeil AC, Shupert WL, Iyasere CA, Hallahan CW, Mican JA, et al. (2001)
High-level HIV-1 viremia suppresses viral antigen-specific CD4(+) T cell
proliferation. Proc Natl Acad Sci U S A 98: 13878–13883.
67. Thimme R, Oldach D, Chang KM, Steiger C, Ray SC, et al. (2001)
Determinants of viral clearance and persistence during acute hepatitis C virus
infection. J Exp Med 194: 1395–1406.
68. Haringer B, Lozza L, Steckel B, Geginat J (2009) Identification and
characterization of IL-10/IFN-gamma-producing effector-like T cells with
regulatory function in human blood. J Exp Med 206: 1009–1017.
69. Boussiotis VA, Tsai EY, Yunis EJ, Thim S, Delgado JC, et al. (2000) IL-10-
producing T cells suppress immune responses in anergic tuberculosis patients.
J Clin Invest 105: 1317–1325.
70. Meiler F, Zumkehr J, Klunker S, Ruckert B, Akdis CA, et al. (2008) In vivo
switch to IL-10-secreting T regulatory cells in high dose allergen exposure. J Exp
Med 205: 2887–2898.
71. O’Garra A, Vieira P (2007) T(H)1 cells control themselves by producing
interleukin-10. Nat Rev Immunol 7: 425–428.
72. Li C, Corraliza I, Langhorne J (1999) A defect in interleukin-10 leads to
enhanced malarial disease in Plasmodium chabaudi chabaudi infection in mice.
Infect Immun 67: 4435–4442.
73. WinklerS, WillheimM, BaierK, SchmidD, Aichelburg A, etal. (1998) Reciprocal
regulation of Th1- and Th2-cytokine-producing T cells during clearance of
parasitemia in Plasmodium falciparum malaria. Infect Immun 66: 6040–6044.
74. Flanagan KL, Plebanski M, Odhiambo K, Sheu E, Mwangi T, et al. (2006)
Cellular reactivity to the p. Falciparum protein trap in adult kenyans: novel
epitopes, complex cytokine patterns, and the impact of natural antigenic
variation. Am J Trop Med Hyg 74: 367–375.
75. Gitau EN, Tuju J, Stevenson L, Kimani E, Karanja H, et al. (2012) T-cell
responses to the DBLalpha-tag, a short semi-conserved region of the Plasmodium
falciparum membrane erythrocyte protein 1. PLoS ONE 7: e30095.
76. Roetynck S, Olotu A, Simam J, Marsh K, Stockinger B, et al. (2013) Phenotypic
and functional profiling of CD4 T cell compartment in distinct populations of
healthy adults with different antigenic exposure. PLoS ONE 8: e55195.
77. Freitas do Rosario AP, Langhorne J (2012) T cell-derived IL-10 and its impact
on the regulation of host responses during malaria. International Journal for
Parasitology 42: 549–555.
78. Metenou S, Dembele B, Konate S, Dolo H, Coulibaly YI, et al. (2011) Filarial
infection suppresses malaria-specific multifunctional Th1 and Th17 responses in
malaria and filarial coinfections. Journal of immunology 186: 4725–4733.
79. Brustoski K, Moller U, Kramer M, Petelski A, Brenner S, et al. (2005) IFN-
gamma and IL-10 mediate parasite-specific immune responses of cord blood
cells induced by pregnancy-associated Plasmodium falciparum malaria. Journal
of immunology 174: 1738–1745.
80. Brockman MA, Kwon DS, Tighe DP, Pavlik DF, Rosato PC, et al. (2009) IL-10
is up-regulated in multiple cell types during viremic HIV infection and reversibly
inhibits virus-specific T cells. Blood 114: 346–356.
81. Brooks DG, Trifilo MJ, Edelmann KH, Teyton L, McGavern DB, et al. (2006)
Interleukin-10 determines viral clearance or persistence in vivo. Nat Med 12:
82. Depinay N, Franetich JF, Gruner AC, Mauduit M, Chavatte JM, et al. (2011)
Inhibitory effect of TNF-alpha on malaria pre-erythrocytic stage development:
influence of host hepatocyte/parasite combinations. PLoS ONE 6: e17464.
83. Nussler A, Pied S, Goma J, Renia L, Miltgen F, et al. (1991) TNF inhibits
malaria hepatic stages in vitro via synthesis of IL-6. International Immunology 3:
84. Katrak S, Gasasira A, Arinaitwe E, Kakuru A, Wanzira H, et al. (2009) Safety
and tolerability of artemether-lumefantrine versus dihydroartemisinin-pipera-
quine for malaria in young HIV-infected and uninfected children. Malaria
Journal 8: 272.
85. Sandison TG, Homsy J, Arinaitwe E, Wanzira H, Kakuru A, et al. (2011)
Protective efficacy of co-trimoxazole prophylaxis against malaria in HIV
exposed children in rural Uganda: a randomised clinical trial. BMJ 342: d1617.
86. Vora N, Homsy J, Kakuru A, Arinaitwe E, Wanzira H, et al. (2010)
Breastfeeding and the risk of malaria in children born to HIV-infected and
uninfected mothers in rural Uganda. Journal of Acquired Immune Deficiency
Syndromes 55: 253–261.
87. Arinaitwe E, Sandison TG, Wanzira H, Kakuru A, Homsy J, et al. (2009)
Artemether-lumefantrine versus dihydroartemisinin-piperaquine for falciparum
malaria: a longitudinal, randomized trial in young Ugandan children. Clinical
infectious diseases : an official publication of the Infectious Diseases Society of
America 49: 1629–1637.
88. (2010) Uganda Clinical Guidelines 2010. National Guidelines on Management
of Common Conditions. Kampala, Uganda: Uganda Ministry of Health.
CD4 Response to Malaria in Highly Exposed Children
PLOS Pathogens | www.plospathogens.org 13 January 2014 | Volume 10 | Issue 1 | e1003864
89. Wipasa J, Okell L, Sakkhachornphop S, Suphavilai C, Chawansuntati K, et al.
(2011) Short-lived IFN-gamma effector responses, but long-lived IL-10 memory
responses, to malaria in an area of low malaria endemicity. PLoS pathogens 7:
90. Horowitz A, Newman KC, Evans JH, Korbel DS, Davis DM, et al. (2010)
Cross-talk between T cells and NK cells generates rapid effector responses to
Plasmodium falciparum-infected erythrocytes. J Immunol 184: 6043–6052.
91. Maecker HT, Trotter J (2006) Flow cytometry controls, instrument setup, and
the determination of positivity. Cytometry Part A 69: 1037–1042.
92. Lamoreaux L, Roederer M, Koup R (2006) Intracellular cytokine optimization
and standard operating procedure. Nat Protoc 1: 1507–1516.
93. McLaughlin BE, Baumgarth N, Bigos M, Roederer M, De Rosa SC, et al. (2008)
Nine-color flow cytometry for accurate measurement of T cell subsets and
cytokine responses. Part I: Panel design by an empiric approach. Cytometry
Part A 73: 400–410.
94. Roederer M, Nozzi JL, Nason MX (2011) SPICE: Exploration and analysis of
post-cytometric complex multivariate datasets. Cytometry Part A [Epub ahead
CD4 Response to Malaria in Highly Exposed Children
PLOS Pathogens | www.plospathogens.org14 January 2014 | Volume 10 | Issue 1 | e1003864