Growth Factor Variation in Two Types of Autologous Platelet Biomaterials: PRP Versus PRF.

Autologous platelet biomaterials represent a key source of cytokines and growth factors extensively used for clinical and surgical applications involving tissue regeneration; wound healing and tissue repair. In this communication we discuss the growth factors released by activated platelet rich plasma (PRP) and platelet rich fibrin (PRF) releasate. Our study highlights that significantly higher growth factors (TGF-ß1) are released by activated PRP as compared to releasate of PRF. The various growth factors released by both platelet products are significantly higher than the baseline concentration in the whole blood and have different bio-mechanism hence should be individualized as per the clinical indication.

Role of signal-to-cut-off ratios of anti-hepatitis C virus antibody by enzyme immunoassays along with ID-NAT for screening of whole blood donors in India.

BACKGROUND: The use of elevated signal-to-cut off ratios (S/CO) as an alternate to further supplemental testing (i.e., RIBA) has been included in the guidelines provided by the Centres for Disease Control and Prevention for HCV diagnostic purposes since 2003. With availability of screening by NAT and non availability of RIBA, further confirmation of HCV infection has been possible at the molecular level (RNA). AIMS: To study the role of S/CO ratios of anti hepatitis C virus antibody detection by enzyme immunoassays (EIA) along with ID-NAT for screening of whole blood donors. METHODS: In this study we reviewed the donor screening status for anti HCV from January 2013 to May 2014. All the donations were screened for anti HCV with fourth generation ELISA (BioRad Monolisa Ag-Ab Ultra) as well as with ID NAT (Procleix Ultrio). The S/CO ratio of all the anti-HCV reactive samples were analysed for their presence of HCV RNA. RESULTS: On screening 21,115 donors for HCV, 83 donors (0.39%) were found reactive on pilot tube and repeat plasma bag testing (S/Co ratio ≥1) by ELISA. 41 donors were HCV RNA reactive with ID-NAT. 4 samples out of 41 were NAT yields and 37 were concordant reactive with ELISA. The S/Co ratio of anti-HCV reactive samples ranged from 0.9-11.1 [mean = 5.1; SD ± 2.9] whereas S/Co ratio of anti HCV and NAT reactive samples (concordant positives) ranged from 4.1-11.1 [mean 7.3]. In our analysis we found that S/CO ratio of 4 showed positive predictive value (PPV) and sensitivity of 100%. SUMMARY/CONCLUSIONS: Our study showed that S/CO of 4 for anti HCV on ELISA would have maximum positive predictive value of having donor with HCV RNA. S/CO ratio of 4 is very close to 3.8 which was the CDC guideline. The presence of anti-HCV does not distinguish between current or past infections but a confirmed anti-HCV-positive result indicates the need for counseling and medical evaluation for HCV infection.

Quantification of platelets and platelet derived growth factors from platelet-rich-plasma (PRP) prepared at different centrifugal force (g) and time.

INTRODUCTION: Platelet derived biomaterials represent a key source of cytokines and growth factors extensively used for tissue regeneration; wound healing and tissue repair. Our study was to quantify platelets and growth factors released by PRP when prepared at different centrifugal force (g) and time. MATERIAL AND METHODS: Our study was approved by the institutional ethical committee. One hundred millilitres of whole blood (WB) was collected in bag with CPDA as the anticoagulant(AC); (14 mL for 100 mL WB ratio). Nine aliquots of 10 mL each were made from the bag and set of three aliquots were made a group. PRP was prepared at varying centrifugal force (group A: -110 g, group B: -208 g & group C: -440 g) & time (1: -5 min, 2: -10 min & 3: -20 min). Contents of each PRP prepared were analysed. Commercial sandwich ELISA kits were used to quantify the concentrations of CD62P (Diaclone SAS; France), Platelet derived growth factors-AB (Qayee-Bio; China), transforming growth factor-ß1 (DRG; Germany) and vascular endothelial growth factor (Boster Immuno Leader; USA) released in each PRP prepared. RESULTS: Eight volunteers were enrolled in the study (24-30 years). The baseline blood counts of all the volunteers were comparable (p ≥ 0.05). Mean ± SD of platelet yield of all nine groups ranged from 17.2 ± 4.2% to 78.7 ± 5.7%. Each PRP was activated with calcified thromboplastin to quantify the growth factors released by them. Significantly higher (p < 0.05) transforming growth factor-ß1 and vascular endothelial growth factor were released compared to the baseline. CONCLUSION: Our study highlights the variation in both force (g) and time results in changes at cellular level and growth factor concentrations.

Blood donor notification and counseling: Our experience from a tertiary care hospital in India.

AIMS: To evaluate the response rate of transfusion-transmissible infection (TTI)-reactive donors after notification of their abnormal test results for the year 2012. MATERIALS AND METHODS: This study is an observational descriptive study performed in our department over a period of 1 year. We evaluated the response rate of TTI-reactive donors after notification of their abnormal test results over 1 year as per the existing strategy (three telephonic and two postal communications). RESULTS: During the study period, among the annual donation of 15,322 units, 464 blood donors were found to be seroreactive. Of these 464 seroreactive cases, 47 were HIV positive, 284 were reactive for Hepatitis B surface antigen (HBsAg), 49 were Hepatitis C (HCV) positive and 84 were VDRL reactive. The TTI-reactive donors (464) for various markers were contacted: 229 (49.4%) telephonically and the remaining 235 (50.6%) not contacted on phone were informed by post. Of the 229 contacted donors, the response rate was 98.2% as only 225 donors reported (221 on the first, three on second and one on the third call) for one to one counseling. The remaining four non-responders were - one HIV and three HBsAg reactive. The remaining 235 (50.6%) reactive donors did not respond to any communication. CONCLUSION: Donor notification and post-donation counseling are an essential aspect of the blood bank that entails provision of information on serological status, assess the impact of test results on the donor and finally referral for medical care. As in our data only 49.4% of the blood donors could be contacted successfully, incomplete demographic details was the major limiting factor in communicating with rest. Of the 229 contacted donors, the response rate was 98.2%. A large majority (94.75%) of the notified donors in our study contacted their health care provider when given clear instructions to do so. These results are encouraging because they indicate that a major element of the notification message is acted upon when it is worded clearly. The very high response rate of the contacted donors ensured their concern for knowing their test result status.

Maternal anti-M induced hemolytic disease of newborn followed by prolonged anemia in newborn twins.

Allo-anti-M often has an immunoglobulin G (IgG) component but is rarely clinically significant. We report a case of hemolytic disease of the fetus and newborn along with prolonged anemia in newborn twins that persisted for up to 70 days postbirth. The aim was to diagnose and successfully manage hemolytic disease of newborn (HDN) due to maternal alloimmunization. Direct antiglobulin test (DAT), antigen typing, irregular antibody screening and identification were done by polyspecific antihuman globulin cards and standard tube method. At presentation, the newborn twins (T1, T2) had HDN with resultant low reticulocyte count and prolonged anemia, which continued for up to 70 days of life. Blood group of the twins and the mother was O RhD positive. DAT of the both newborns at birth was negative. Anti-M was detected in mothers as well as newborns. Type of antibody in mother was IgG and IgM type whereas in twins it was IgG type only. M antigen negative blood was transfused thrice to twin-1 and twice to twin-2. Recurring reduction of the hematocrit along with low reticulocyte count and normal other cell line indicated a pure red cell aplastic state. Anti-M is capable of causing HDN as well as prolonged anemia (red cell aplasia) due to its ability to destroy the erythroid precursor cells. Newborns with anemia should be evaluated for all the possible causes to establish a diagnosis and its efficient management. Mother should be closely monitored for future pregnancies as well.

Sensitivity of individual donor nucleic acid testing (NAT) for the detection of hepatitis B infection by studying diluted NAT yield samples.

BACKGROUND: Screening blood donors for the presence of hepatitis B virus surface antigen (HBsAg) has been the backbone of blood safety. However, occult hepatitis B infection (OBI) in donors can be missed when only HBsAg screening is used. Nucleic acid testing (NAT) is capable of detecting OBI among donors. The aim of our study was to analyse the sensitivity of NAT for detecting OBI. MATERIAL AND METHODS: The kits used during the study for serology testing were BioRad Monolisa™ HBsAg Ultra (HBsAg screening), Abbott Architect for anti-HBcAg (total) and anti-HBsAg testing, and Vitros® by Ortho Clinical Diagnostics for anti-HBcAg (IgM). Procleix Ultrio was used for individual donor-NAT (ID-NAT) and Abbott m2000 for estimation of HBV DNA. Out of 28,134 HBsAg non-reactive donors, 25 were ID-NAT-reactive. Of these 25 NAT yield samples, 18 were studied further at different dilutions from 1:2 to 1:16. The doubling dilutions were made with HBV non-reactive AB plasma. Undiluted samples were used for all serological tests and for HBV DNA estimation. RESULTS: Of the 18 samples studied, nine were NAT-reactive at a dilution of <1:4 and five out of these showed presence of antibody to core antigen (IgG+IgM). Antibody to surface antigen was present in only two of the nine NAT-reactive samples, one with antibody to core antigen and the other without. Six had a viral load in the range from <10 to 38 IU/mL whereas the viral load in the remaining three samples was not determined. Among the other nine samples which were NAT-reactive at dilutions≥1:4, antibody to core antigen (IgG+IgM) was present in seven. DISCUSSION: Our study showed that ID-NAT testing along with HBsAg screening could detect most potentially HBV infectious donors (including those with OBI). NAT screening for HBV on diluted samples could compromise blood safety because samples with a low viral load will escape detection.

Reactive donor notification: First error reported.

Donor notification and post-donation counseling is an essential role of blood bank. If a donor is reactive for any marker, the blood bank counselor, informs the donor and advices him/her to report to the blood bank for further counseling and management. The counselor at our blood bank informed a young female voluntary donor to be reactive for HIV both with ELISA as well as NAT. When the donor reported to blood bank, the repeat testing was negative and no history of high risk behavior could be elicited. The hospital information system (HIS) records were checked again immediately for clarification and showed consistency with her demographic profile. But when her manual records and donor questionnaire were retrieved, showed information displayed in the HIS system was wrongly interpreted by the counselor. In this era of information technology being highly advanced, the role of manual record keeping is still the gold standard.

Serological characterization of occult hepatitis B virus infection among blood donors in India.

INTRODUCTION: Discovery of hepatitis B infections characterized by the presence of viral genome without detectable HBsAg (Occult Hepatitis; OBI) has initiated a considerable amount of research in this regard. Our study is a serological and molecular characterization of OBI among the donors who donated at our blood bank during the study period. MATERIAL AND METHOD: During the study period HBsAg ELISA non reactive ID-NAT reactive donors samples were screened for presence of antibody against HBc, HBs and HBe. Molecular analysis of these NAT yield samples was undertaken for detection of the viral load and genotyping. RESULT: We studied 28,134 HBsAg ELISA non reactive donor samples. On testing them with ID-NAT, HBV DNA was detected in 25 samples. Eighteen samples out of these 25 NAT yield were further worked up. The 66.6% of the NAT yield samples (12 out of 18) were reactive for antibody against HBc. The 25% (3 out of 12) of these NAT yield samples having antibody against core antigen also had antibody against HBs. The 27.7% (5 out of 18) of NAT yield detected by ID-NAT did not have any detectable serological marker in blood. Four out of 12 core antibody positive NAT yield samples had genotype A HBV infection. CONCLUSION: As per our study molecular detection of HBV DNA by ID-NAT, we were able to analyze 18 HBV NAT yield cases among 28,134 HBsAg ELISA non reactive donors. Out of 18, 12 donors were OBI whereas the rest (6) were in window period (WP yield) of HBV infection. One out of every 3.6 NAT yield detected by ID-NAT was non reactive for all serological markers.