Analysis of maternal Fc gamma receptor IIIb isoantibodies using immunomagnetic negative selected neutrophils.

BACKGROUND AND OBJECTIVES: The isolation of neutrophils and subsequent detection of anti-human neutrophil antigens (HNA) antibodies are crucial in clinical medicine for the diagnosis of autoimmune neutropenia, neonatal alloimmune neutropenia (NAIN) and transfusion-related acute lung injury (TRALI). This study reports two cases of maternal anti-Fc-gamma-receptor-IIIb (FcγRIIIb) isoimmunization without NAIN symptoms and compares the efficiency of immunomagnetic negative selection (IMNS) with traditional dextran/Ficoll for neutrophil isolation in HNA serological assays. MATERIALS AND METHODS: Investigating two cases of maternal anti-FcγRIIIb isoimmunization, neutrophils from three donors were isolated from 8 mL of whole blood using IMNS and dextran/Ficoll. Serological assays included the granulocyte agglutination and immunofluorescence test, monoclonal antibody immobilization of granulocyte antigens and the LABScreen Multi (One Lambda). IMNS and dextran/Ficoll were compared in terms of cell yield, viability, time, cost and purity. RESULTS: Maternal anti-FcγRIIIb isoantibodies with FCGR3B gene deletion were detected in both cases. Newborns and fathers exhibited specific gene combinations: FCGR3B*02/FCGR3B*02 (Case 1) and FCGR3B*02/FCGR3B*03 (Case 2). IMNS outperformed dextran/Ficoll, yielding four times more neutrophils (average neutrophil counts: 18.5 × 103/µL vs. 4.5 × 103/µL), efficiently removing non-neutrophil cells and reducing processing time (30-40 min vs. 70-90 min), although it incurred a higher cost (2.7 times). CONCLUSION: Two cases of maternal anti-FcγRIIIb isoantibodies, unrelated to NAIN, were identified. Although neutropenia has not been described in these cases, we emphasize the importance of identifying asymptomatic cases with the potential for severe neutropenia. Additionally, IMNS is introduced as a rapid, high-yield, high-purity neutrophil isolation technique, beneficial for serological assays detecting anti-HNA antibodies.

Antibodies to human neutrophil antigen HNA-3b implicated in cases of neonatal alloimmune neutropenia.

BACKGROUND: Neonatal alloimmune neutropenia results from maternal alloimmunization to human neutrophil antigens. The alloantibodies involved in neonatal alloimmune neutropenia are against human neutrophil antigens HNA-1a, HNA-1b, HNA-1c, HNA-1d, HNA-2, HNA-3a, HNA-4a, HNA-4b, and HNA-5a; however, to date, antibodies specific to HNA-3b have not been reported. STUDY DESIGN AND METHODS: Blood samples from 10,000 unselected neonates were analyzed, resulting in the selection of 88 neutropenic newborns (neutrophil count <1.5 × 109 /L) from 83 mothers (three pairs of twins and one triplet). HNA-3 genotyping was performed by polymerase chain reaction-restriction fragment length polymorphism to identify the cases of maternal-fetal HNA-3 incompatibility. Serologic studies for detecting maternal HNA-3 alloantibodies were performed with the granulocyte agglutination test, the white blood cell immunofluorescence test, and a LABScreen Multi-HNA Kit. RESULTS: Genotyping studies identified 13 of 88 (14.8%) instances of maternal-fetal HNA-3 incompatibility, with all mothers typed as HNA-3a/a and neonates typed as HNA-3a/b. Serologic studies revealed that five of 13 (38.5%) mothers carried anti-HNA-3b plus human leukocyte antigen antibodies and that three of 13 (23.1%) mothers had anti-HNA-3b without human leukocyte antigen antibodies. CONCLUSION: Here, we report the first three cases of neonatal alloimmune neutropenia associated with HNA-3b antibodies resulting in a neonatal alloimmune neutropenia incidence of one in 3333 live births.

Direct antiglobulin test in the differential diagnosis of ABO hemolytic disease of the newborn: an important tool with high negative predictive value.

BACKGROUND: Hemolysis due to ABO incompatibility is an important differential diagnosis in newborns presenting with jaundice. Clinical studies evaluating ABO hemolytic disease of fetus and newborn (ABO-HDFN) question the diagnostic value of the direct antiglobulin test (DAT) in this situation. GOALS: To determine the clinical and laboratorial findings associated with the occurrence of ABO-HDFN and to evaluate the accuracy of DAT as a diagnostic tool. METHODS: This was a nested case control study with a cohort of 4122 newborns. Clinical and immunohematological data were retrieved from medical files including clinical and laboratorial factors associated with ABO-HDFN. Sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) of positive DAT were calculated. RESULTS: Among the 4122 newborns, 44 had the diagnosis of ABO-HDFN. Positive DAT, group O mother and group A newborn were significantly associated with the occurrence of neonatal jaundice and this association persisted in a multivariable model (p-value <0.001). DAT presented 65.85 % sensitivity, 96.28 % specificity, 16.9 % PPV and 99.6 % NPV for the diagnosis of ABO-HDFN. There were no cases of positive DAT in cases other than O/A and O/B incompatibilities. The newborn hemoglobin was significantly lower in O/A incompatibility (p-value <0.001). CONCLUSION: Positive DAT, mother of group O and newborn of group A are independent risk factors associated with ABO-HDFN. DAT exhibited high NPV for the diagnosis of this complication. Thus, performing DAT in newborns with O/A and O/B incompatibilities is a cost-effective strategy that can be applied as routine by blood banks.

INCIDENCE OF SMALL FOR GESTATIONAL AGE NEONATES, ACCORDING TO THE FENTON AND INTERGROWTH-21ST CURVES IN A LEVEL II MATERNITY.

OBJECTIVE: To compare the incidence of small for gestational age infants among late preterm and term newborns, using the Fenton and Intergrowth-21st curves. METHODS: Observational and retrospective study with newborns in a level II maternity. The study was approved by the Institution's Ethics Committee. Live births from July 2007 to February 2009 with a gestational age from 34 to 41 weeks and seven days were included. Neonates with incomplete data were excluded. Appropriate weight for gestational age was assessed by the Fenton and Intergrowth-21st intrauterine growth curves, considering birth weight <10th percentile as small for gestational age. The degree of agreement between the two curves was assessed by the Kappa coefficient. Numerical variables were compared using the Student t-test or the Mann-Whitney. Categorical variables were compared using the chi-square test. Statistical analyzes were performed using SPSS17® software, considering significant, p<0.05. RESULTS: We included 2849 newborns with a birthweight of 3210±483 g, gestational age of 38.8±1.4 weeks; 51.1% male. The incidence of small for gestational age in the full sample was 13.0 vs. 8.7% (p<0.001, Kappa=0.667) by the Fenton and Intergrowth-21st curves, respectively. Among late preterm, the incidence of small neonates was 11.3 vs. 10.9% (p<0.001; Kappa=0.793) and among full-term infants it was 13.1% vs. 8.5% (p<0.001; Kappa=0.656), respectively for the Fenton and Intergrowth-21st curves. CONCLUSIONS: The incidence of small for gestational age newborns was significantly higher using the Fenton curve, with greater agreement between the Fenton and Intergrowth-21st curves among late preterm, compared to full term neonates.

BLOOD CELLS PROFILE IN UMBILICAL CORD OF LATE PRETERM AND TERM NEWBORNS.

OBJECTIVE: To describe the hematological profile in cord blood of late preterm and term newborns and compare blood indices according to sex, weight for gestational age and type of delivery. METHODS: Cross-sectional study with late preterm and term newborns in a second-level maternity. Multiple gestation, chorioamnionitis, maternal or fetal hemorrhage, suspected congenital infection, 5-minute Apgar <6, congenital malformations, and Rh hemolytic disease were excluded. Percentiles 3, 5,10, 25, 50, 75, 90, 95 and 97 of blood indices were calculated for both groups. RESULTS: 2,662 newborns were included in the sample, 51.1% males, 7.3% late preterms, 7.8% small for gestational age (SGA) and 81.2% adequate for gestational age (AGA). Mean gestational age was 35.6±1.9 and 39.3±1.0 weeks, respectively, for premature and term neonates. The erythrocytes indices and white blood cells increased from 34-36.9 to 37-41.9 weeks. Basophils and platelets remained constant during gestation. Premature neonates presented lower values ​​of all blood cells, except for lymphocytes and eosinophils. SGA neonates presented higher values ​​of hemoglobin, hematocrit and lower values of leukocytes, neutrophils, bands, segmented, eosinophils, monocytes and platelets. Male neonates presented similar values ​​of erythrocytes and hemoglobin and lower leukocytes, neutrophils, segmented and platelets. Neonates delivered by C-section had lower values ​​of red blood cells and platelets. Chronic or gestational hypertension induced lower number of platelets. CONCLUSIONS: Blood cells increased during gestation, except for platelets and basophils. SGA neonates had higher hemoglobin and hematocrit values and lower leukocytes. Number of platelets was smaller in male SGAs, born by C-section and whose mothers had hypertension.

Incidence of small for gestational age neonates, according to the fenton and intergrowth-21st curves in a level ii maternity / Incidência de recém-nascidos pequenos para a idade gestacional segundo curva de fenton e intergrowth-21st em uma maternidade de nível secundário

ABSTRACT Objective: To compare the incidence of small for gestational age infants among late preterm and term newborns, using the Fenton and Intergrowth-21st curves. Methods: Observational and retrospective study with newborns in a level II maternity. The study was approved by the Institution's Ethics Committee. Live births from July 2007 to February 2009 with a gestational age from 34 to 41 weeks and seven days were included. Neonates with incomplete data were excluded. Appropriate weight for gestational age was assessed by the Fenton and Intergrowth-21st intrauterine growth curves, considering birth weight <10th percentile as small for gestational age. The degree of agreement between the two curves was assessed by the Kappa coefficient. Numerical variables were compared using the Student t-test or the Mann-Whitney. Categorical variables were compared using the chi-square test. Statistical analyzes were performed using SPSS17® software, considering significant, p<0.05. Results: We included 2849 newborns with a birthweight of 3210±483 g, gestational age of 38.8±1.4 weeks; 51.1% male. The incidence of small for gestational age in the full sample was 13.0 vs. 8.7% (p<0.001, Kappa=0.667) by the Fenton and Intergrowth-21st curves, respectively. Among late preterm, the incidence of small neonates was 11.3 vs. 10.9% (p<0.001; Kappa=0.793) and among full-term infants it was 13.1% vs. 8.5% (p<0.001; Kappa=0.656), respectively for the Fenton and Intergrowth-21st curves. Conclusions: The incidence of small for gestational age newborns was significantly higher using the Fenton curve, with greater agreement between the Fenton and Intergrowth-21st curves among late preterm, compared to full term neonates.

RESUMO Objetivo: Comparar a incidência de neonatos pequenos para idade gestacional entre nascidos vivos pré-termo tardios e a termo utilizando as curvas de Fenton e Intergrowth-21st. Métodos: Estudo observacional retrospectivo com recém-nascidos de uma maternidade pública de nível secundário. Foram incluídos nascidos vivos de julho/2007 a fevereiro/2009 com idade gestacional de 34 a 41 semanas e seis dias. O estudo foi aprovado pelo Comitê de Ética da instituição. Foram excluídos recém-nascidos com dados incompletos. Para adequação do peso/da idade gestacional, utilizaram-se as curvas de crescimento intrauterino de Fenton e Intergrowth-21st, considerando-se pequeno aquele com peso ao nascer <10º percentil. O grau de concordância entre as duas curvas foi avaliado pelo coeficiente Kappa. As variáveis numéricas foram comparadas pelo teste t de Student ou de Mann-Whitney, conforme distribuição, e as categóricas pelo teste χ2. As análises estatísticas foram realizadas no programa Statistical Package for the Social Sciences (SPSS) 17®, considerando-se significante p<0,05. Resultados: Foram incluídos 2.849 recém-nascidos com peso ao nascer de 3210±483 g, idade gestacional de 38,8±1,4 semanas, sendo 51,1% masculinos. A incidência de recém-nascidos pequenos para a idade gestacional pela curva de Fenton e Intergrowth-21st na amostra total foi, respectivamente, de 13 e 8,7% (p<0,001; Kappa=0,667). Entre os pré-termo tardios, a incidência foi de 11,3 e 10,9% (p<0,001; Kappa=0,793) e entre os nascidos a termo foi de 13,1 e 8,5%, (p<0,001; Kappa=0,656), respectivamente, para as curvas de Fenton e Intergrowth-21st. Conclusões: A incidência de recém-nascidos pequenos para idade gestacional foi significantemente maior pela curva de Fenton, com maior concordância entre as curvas de Fenton e Intergrowth-21st em recém-nascidos pré-termo tardios do que nos nascidos a termo.

Blood cells profile in umbilical cord of late preterm and term newborns / Perfil de hemograma em sangue de cordão umbilical de recém-nascidos pré-termo tardio e a termo

ABSTRACT Objective: To describe the hematological profile in cord blood of late preterm and term newborns and compare blood indices according to sex, weight for gestational age and type of delivery. Methods: Cross-sectional study with late preterm and term newborns in a second-level maternity. Multiple gestation, chorioamnionitis, maternal or fetal hemorrhage, suspected congenital infection, 5-minute Apgar <6, congenital malformations, and Rh hemolytic disease were excluded. Percentiles 3, 5,10, 25, 50, 75, 90, 95 and 97 of blood indices were calculated for both groups. Results: 2,662 newborns were included in the sample, 51.1% males, 7.3% late preterms, 7.8% small for gestational age (SGA) and 81.2% adequate for gestational age (AGA). Mean gestational age was 35.6±1.9 and 39.3±1.0 weeks, respectively, for premature and term neonates. The erythrocytes indices and white blood cells increased from 34-36.9 to 37-41.9 weeks. Basophils and platelets remained constant during gestation. Premature neonates presented lower values ​​of all blood cells, except for lymphocytes and eosinophils. SGA neonates presented higher values ​​of hemoglobin, hematocrit and lower values of leukocytes, neutrophils, bands, segmented, eosinophils, monocytes and platelets. Male neonates presented similar values ​​of erythrocytes and hemoglobin and lower leukocytes, neutrophils, segmented and platelets. Neonates delivered by C-section had lower values ​​of red blood cells and platelets. Chronic or gestational hypertension induced lower number of platelets. Conclusions: Blood cells increased during gestation, except for platelets and basophils. SGA neonates had higher hemoglobin and hematocrit values and lower leukocytes. Number of platelets was smaller in male SGAs, born by C-section and whose mothers had hypertension.

RESUMO Objetivo: Descrever o perfil hematológico em sangue de cordão de recém-nascidos pré-termo tardio e a termo e comparar parâmetros hematimétricos segundo sexo, adequação peso idade gestacional e tipo de parto. Métodos: Estudo transversal com recém-nascidos pré-termo tardio e a termo, em maternidade de nível secundário. Excluíram-se gestação múltipla, corioamnionite, hemorragia materna ou fetal, suspeita de infecção congênita, Apgar no 5o minuto <6, malformações congênitas e doença hemolítica Rh. Calcularam-se os percentis 3, 5, 10, 25, 50, 75, 90, 95 e 97 dos parâmetros hematológicos. Resultados: Incluíram-se 2.662 recém-nascidos, 51,1% do sexo masculino, 7,3% prematuros tardios, 7,8% pequenos para a idade gestacional e 81,2% adequados. A idade gestacional foi 35,6±1,9 e 39,3±1,0 semanas, respectivamente, nos prematuros e termos. As séries vermelha e branca aumentaram de 34-36,9 para 37-41,9 semanas, exceto basófilos e plaquetas, que permaneceram constantes. Os prematuros apresentaram menores médias nas séries vermelha, plaquetária e branca, com exceção de linfócitos e eosinófilos. Recém-nascidos pequenos para a idade gestacional apresentaram maiores valores de hemoglobina e hematócrito e menores de leucócitos, neutrófilos, bastonetes segmentados, eosinófilos, monócitos e plaquetas. Recém-nascidos masculinos apresentaram taxas semelhantes de hemoglobina e hematócrito e menores de leucócitos, neutrófilos, segmentados e plaquetas. Na cesárea, as células vermelhas e as plaquetas foram menores que no parto vaginal. O número de plaquetas foi menor na hipertensão crônica ou gestacional. Conclusões: As células sanguíneas aumentaram durante a gestação, exceto plaquetas e basófilos. Recém-nascidos pequenos para a idade gestacional apresentaram maiores taxas de hemoglobina e hematócrito e menores de células brancas. O número de plaquetas foi menor no recém-nascido pequeno para a idade gestacional, masculino, nascido por cesárea e de mãe hipertensa.