Comparison of seven heterophile antibody assays for laboratory diagnosis of infectious mononucleosis in pediatric patients.

Heterophile antibody assays have been used to aid the diagnosis of infectious mononucleosis caused by the Epstein-Barr virus. Seven commercially available assays currently widely utilized in clinical laboratories were compared in this study. Variable performance characteristics and assay times are observed, and these pieces of data may assist clinical laboratories in assay selection and result interpretation.

Aspects of histocompatibility testing in xenotransplantation.

Xenotransplantation, using genetically-modified pigs for clinical organ transplantation, is a solution to the organ shortage. The biggest barrier to clinical implementation is the antigenicity of pig cells. Humans possess preformed antibody to pig cells that initiate antibody-mediated rejection of pig organs in primates. Advances in genetic engineering have led to the development of a pig lacking the three known glycan xenoantigens (triple-knockout [TKO] pigs). A significant number of human sera demonstrate no antibody binding to TKO pig cells. As a result of the TKO pig's low antigen expression, survival of life-supporting pig organs in immunosuppressed nonhuman primates has significantly increased, and hope has been renewed for clinical trials of xenotransplantation. It is important to understand the context in which xenotransplantation's predecessor, allotransplantation, has been successful, and the steps needed for the success of xenotransplantation. Successful allotransplantation has been based on two main immunological approaches - (i) adequate immunosuppressive therapy, and (ii) careful histocompatibility matching. In vivo studies suggest that the available immunosuppressive regimens are adequate to suppress the human anti-pig cellular response. Methods to evaluate and screen patients for the first clinical xenotransplantation trial are the next challenge. The goal of this review is to summarize the history of histocompatibility testing, and the available tools that can be utilized to determine xenograft histocompatibility.

Prevalence of heterophilic antibodies in serum samples from horses in an equine hospital, and elimination of interference using chicken IgY.

BACKGROUND: Heterophilic antibodies in serum and plasma can interfere with mammalian antibodies in immunoassays and result in false test results, usually false positive. Although studies screening for heterophilic antibodies as well as elimination studies have been conducted in dogs and cats, knowledge of the presence of heterophilic antibodies in other species in veterinary medicine is limited. In this study, a 2-site sandwich-type interference assay that detects anti-mouse antibodies was used to detect heterophilic antibodies in a population of horses treated in an animal hospital. RESULTS: A total of 194 serum samples from 127 individual horses were analyzed. There were 11/127 (8.7%) interference-positive horses, and these were analyzed in an assay exchanging the capture mouse IgG with chicken IgY. The positive samples were negative in the chicken IgY assay, indicating elimination of a possible interference, with the chicken-based assay. Four interference-positive samples were from geldings, and anti-Müllerian hormone (AMH) was analyzed from these samples. AMH concentrations were negative in these samples as expected in geldings, indicating that the heterophilic antibodies did not cause interference in the AMH assay. CONCLUSION: The present study shows that there are heterophilic antibodies in horse serum samples like in samples from humans, dogs, and cats. The use of chicken-based reagents, such as chicken IgY, which do not cross-react with mammalian IgG, eliminates the effects of interfering antibodies in the samples. Equine heterophilic antibodies do not necessarily cause interference in commercial immunoassays.

The Role of SLAs in Xenotransplantation.

Advances in genetic engineering, particularly CRISPR/Cas9, have resulted in the development of a triple glycan-knockout (TKO) pig. There is minimal human antipig antibody binding to TKO pig cells. The TKO background has decreased antibody binding to a sufficiently low level that any additional xenoantigens expressed on the cells can now be more easily detected. One of these xenoantigens is the swine major histocompatibility complex, termed swine leukocyte antigens (SLA). SLA are the homolog to HLAs, a protein complex expressed on human tissue capable of stimulating the development of new antibodies in allotransplantation. These antibodies can result in graft failure through hyperacute, acute, or chronic rejection. Our knowledge of SLA, particularly in the last 5 years, has grown considerably. The presence, cause, and methods to detect anti-SLA antibodies will need to be carefully considered for the first clinical trial of xenotransplantation. The focus of this review is to summarize the role of SLA in xenotransplantation and consider whether it will prove to be a major barrier. Techniques are now available to mutate target SLA amino acids to ensure that cross-reactive anti-HLA antibodies no longer bind to SLA on the cells of the organ-source pigs. While deletion of SLA expression is possible, it would render the pig at risk for infectious complications. The ideal organ-source pig for HLA highly sensitized recipients may therefore be 1 with site-specific mutations to eliminate cross-reactive binding.

Anti-pig IgE and IgA Antibodies in Naive Primates and Nonhuman Primates With Pig Xenografts.

BACKGROUND: Natural preformed anti-pig IgM/IgG antibodies in primates play an important role in xenograft rejection. As it is not clear how IgE and IgA engage in the immune system in xenotransplantation, we investigated natural preformed and elicited anti-pig IgE/IgA in naive primates and after xenotransplantation in nonhuman primates. METHODS: The binding of IgM/IgG/IgE/IgA antibodies to red blood cells (RBCs) from wild-type (WT), α1,3-galactosyltransferase gene-knockout (GTKO), and GTKO/cytidine monophospho-N-acetylneuraminic acid hydroxylase gene-knockout/ß-1,4 N-acetylgalactosaminyltransferase 2 gene-knockout (ie, triple-knockout pigs) pigs were measured by flow cytometry in naive human (n = 50) and baboon (n = 14) sera. Antibody binding to WT and GTKO pig RBCs (pRBCs) was also measured in the sera of baboons (nonsensitized n = 7, sensitized n = 2) and rhesus monkeys (nonsensitized n = 2, sensitized n = 11) following WT or GTKO pig organ/tissue xenotransplantation. Deposition of IgM/IgG/IgE/IgA in the grafts was detected by immunohistochemistry. RESULTS: The majority of humans had natural preformed IgM/IgG/IgE/IgA to WT and GTKO pRBCs. In contrast, IgM/IgG/IgE/IgA to triple-knockout pRBCs were present at lower levels and frequency (P < 0.01). Baboons also had IgM/IgG/IgE/IgA antibodies against WT pRBCs, but fewer to GTKO and triple-knockout (P < 0.01). After xenotransplantation into nonhuman primates, when IgM/IgG increased, IgE/IgA also increased, but to a lesser extent. In addition to IgM/IgG, IgE or IgA deposition was observed in rejected pig xenografts. CONCLUSIONS: Primates develop serum anti-pig IgE/IgA antibodies both naturally and during xenograft rejection. The pathophysiological role, if any, of anti-pig IgE/IgA antibodies remains unknown.

Broiler stress responses to light intensity, flooring type, and leg weakness as assessed by heterophil-to-lymphocyte ratios, serum corticosterone, infrared thermography, and latency to lie.

Stress and leg weakness are detrimental to broiler production, health, and welfare. Traditional methods to evaluate stress may be stressful to the bird because they are invasive and require handling and restraint. Two studies examined the effects of light intensity and flooring on the following in broilers: 1) traditional methods for assessing stress using heterophil-to-lymphocyte ratios and serum corticosterone (CORT) concentrations, 2) noninvasive measures of stress from infrared thermography (IRT) eye and beak surface temperatures, and 3) latency-to-lie (LTL) test times of birds tested individually and in groups of 5. Day-of-hatch male broiler chicks were placed into 6 pens (N = 120 chicks/pen). At 1 wk, pens were allocated to 3 light intensity treatments (2, 5, or 10 lux). At 4 wk, half of the birds from each pen were moved to a pen with wire flooring and the same light intensity. At 1, 4, 5, and 8 wk, blood samples were collected and IRT images of the heads of 5 clinically healthy broilers from each pen were captured. In study 2, IRT images of the heads of birds that became lame in the wire flooring pens were taken. There were no treatment effects on the LTL times of birds tested in groups or individually (P > 0.05). On day 56 in study 1, birds on wire flooring had elevated heterophil-to-lymphocyte ratios and CORT concentrations (P ≤ 0.002) and depressed IRT eye and beak temperatures (P < 0.0001). In both studies, there were negative correlations between CORT concentrations and IRT beak surface temperatures (P < 0.05). Lame birds had lower IRT eye and beak surface temperatures than sound birds (P ≤ 0.004), and the IRT beak surface temperatures of lame birds were lower than their eye surface temperatures (P = 0.004) in study 2. These studies indicate that the IRT surface temperatures of the eye, and more distinctly of the beak, can be used as sensitive noninvasive indicators of stress.

Relapse of Nephrotic Syndrome after Adrenocorticotropic Hormone-Induced Remission: Implications of Adrenocorticotropic Hormone Antibodies.

BACKGROUND: Prolonged use of corticosteroids continues to be the mainstay in the management of most proteinuric glomerulopathies, but is limited by extensive side effects. Alternative medications such as adrenocorticotropic hormone (ACTH) have been recently used to treat refractory glomerulopathies and have shown superior outcomes when compared with steroids. However, the clinical responsiveness to ACTH therapy varies considerably with a number of patients exhibiting de novo or acquired resistance. The underlying mechanism remains unknown. METHODS: A patient with steroid-dependent focal segmental glomerulosclerosis (FSGS) developed severe steroid side effects impacting quality of life and was converted to repository porcine ACTH therapy. Immediate response in the form of remission of nephrotic syndrome was noted followed by relapse in 10 weeks. Suspecting the role of some ACTH-antagonizing factors, the patient's serum was examined. RESULTS: Immunoblot-based antibody assay revealed high titers of de novo IgG antibodies in the patient's serum that were reactive to the porcine corticotropin with negligible cross-reactivity to human corticotropin. In vitro, in cultured B16 melanoma cells that express abundant melanocortin receptors, addition of the patient's serum substantially abrogated the porcine corticotropin triggered signaling activity of the melanocortinergic pathway, marked by phosphorylation of glycogen synthase kinase 3ß, thus suggesting a mitigating effect on the biological functionality of porcine corticotropin. CONCLUSION: ACTH is a useful alternative therapeutic modality for refractory proteinuric glomerulopathies like FSGS. However, as quintessential therapeutic biologics, natural ACTH, regardless of purity and origin, is inevitably antigenic and may cause the formation of neutralizing antibodies in some sensitive patients, followed by resistance to ACTH therapy. It is imperative to develop ACTH analogues with less immunogenicity for improving its responsiveness in patients with glomerular diseases.

[What to do in the event of a suspicion of analytical interference during an immunoassay?] / Conduite à tenir devant une suspicion d'interférence analytique lors d'un immunodosage.

Identifying analytical interference is a challenge for the medical biologist in providing advice to the prescriber. Indeed, these analytical interferences often have deleterious consequences on the care of patients. Understanding their mechanisms and mastering corrective procedures is essential to limit these management errors. Faced with the many questions from clinicians in current practice, we propose an algorithm for managing a sample when interference is suspected.

Interpretation of EBV serology for human body material donors: Is there a need for early antigen IgG and heterophile antibodies testing?

In this report we evaluated a diagnostic algorithm, proposed by the Belgian Superior Health Council, to detect acute and past Epstein-Barr virus (EBV) infections by means of serology in donors of human body material for transplantation. The available EBV serology parameters were tested on eighty serum samples on three random access analysers: Architect i2000 SR, Liasion XL and BioPlex 2200. The EBV sero-status was determined according to the proposed algorithm and results were compared between the different analysers. Seventy one % of the samples gave concordant interpretations on the three analysers. Most of the discordant results were attributable to early antigen (EA) IgG. The knowledge of the EA IgG and heterophile antibodies (HA) IgM status provided only limited added value and was only useful to distinguish between a very early acute infection and false positivity of viral capsid antigen IgM. The diagnostic algorithm proposed by the Belgian Superior Health Council is merely directive and each individual lab remains responsible for the interpretation and implementation of test combinations for the detection of EBV infections. Our study shows the limited added value of testing for EA IgG and HA IgM, based both on clinical and technical performance.

Human CTLA4-Ig therapy can give false-positive anti-pig antibody results in primates after xenotransplantation.

BACKGROUND: Measurement of serum anti-pig antibodies is an important parameter in immune monitoring after pig-to-nonhuman primate xenotransplantation. Pig aortic endothelial cells (pAECs) are commonly used for this purpose. However, human (h) CTLA4-Ig (abatacept/belatacept) could bind to pCD80/86 on the cells, and a secondary antibody (i.e., anti-human IgG) may recognize hCTLA4-Ig (in the absence of serum anti-pig IgG antibody binding to pAECs), potentially leading to misinterpretation of the results. Our aim was to determine whether hCTLA4-Ig binding to pAECs is associated with false-positive results. METHODS: Sera were obtained from (i) naïve baboons (n = 3) and (ii) baboons (n = 2) that had undergone pig artery patch transplantation with/without hCTLA4Ig therapy. Serum IgM and IgG binding to (i) AECs, (ii) red blood cells (RBCs), and (iii) CD3+T cells in peripheral blood mononuclear cells (PBMCs) from an α1,3-galactosyltransferase gene-knockout pig expressing human CD46 (GTKO/hCD46) was measured by flow cytometry in the presence or absence of hCTLA-4Ig. Complement-dependent cytotoxicity (CDC) of wild-type (WT) pAECs by hCTLA4Ig was measured by flow cytometry. RESULTS: Sera containing hCTLA4-Ig demonstrated significantly increased IgG (but not IgM) binding to pAECs (relative geometric mean [rGM] = 1.8) compared to sera without hCTLA-4Ig (rGM =1.3) (p < .01). In contrast, there was no increased binding to pRBCs or CD3+T cells. hCTLA4-Ig did not result in cytotoxicity of WT pAECs. CONCLUSIONS: pAECs might not be an optimal cell to investigate anti-pig IgG binding when hCTLA4-Ig is administered to the recipient, as a false-positive result may result from hCTLA4-Ig binding to the pAECs. CD3+T cells would be preferable targets (compared to pRBCs) because they express both carbohydrate and MHC class I/II antigens.

PTH Immunoassay Interference Due to Human Anti-Mouse Antibodies in a Subject With Obesity With Normal Parathyroid Function.

CONTEXT: Immunoassay interference has been most often found with prolactin measurement. However, only few data exist on immunoassay interference for other hormones. CASE DESCRIPTION: A 36-year-old woman with obesity (body mass index, 31 kg/m2) had regularly attended our endocrine unit for type 2 diabetes therapy. When she was included as a control subject in a study for obesity management, detailed laboratory testing was performed, including PTH. In the absence of clinical symptoms, she presented with normal calcium, phosphate, and vitamin D levels. However, the PTH levels were >5000 ng/L. These results were obtained using the Roche Elecsys electrochemiluminescence assay. Repeated measurements with this assay (mouse antibody) led to the same findings. However, using an Euroimmun assay (goat antibody), the exact PTH values were measured at 18.0 ng/L. After pretreatment with a heterophilic antibody blocking reagent, the results of the Roche assay had decreased to a normal level. This phenomenon was explained by the detection of human anti-mouse antibodies in the proband's serum. CONCLUSIONS: In cases of prolactin immunoassay interference, endogenous antibodies will bind to the hormone in vivo, resulting in complexes of a high molecular weight that are less efficiently cleared by the kidneys and, thus, accumulate in the blood. In contrast, the PTH values >5000 ng/L detected in our subject most likely had resulted from the specific interference of the human anti-mouse antibodies present in the proband's serum with the assay antibody, resulting in artificial stimulation of the Roche assay detection system ex vivo.

Circulating pig-specific DNA as a novel biomarker for monitoring xenograft rejection.

Monitoring for immune rejection is crucial for long-term survival of pig xenografts. Circulating DNA is a promising non-invasive biomarker for either organ injury or response to therapy. In this study, circulating pig-specific DNA (cpsDNA) was monitored during xenograft rejection. Potential targets of cpsDNA were selected by in silico analysis, and species specificity of selected primers was confirmed by PCR. Subsequently, cpsDNA as a biomarker was evaluated using a complement-dependent cytotoxicity (CDC) assay in vitro. Then, early diagnosis and response to rapamycin were assessed by an in vivo imaging model of pig-to-mouse cell transplantation. Finally, cpsDNA was monitored in a pig-to-monkey artery patch transplantation model. The results showed that (a) a method of cpsDNA quantitation was established for application in mouse and nonhuman primate models; (b) cpsDNA reflected CDC in vitro; (c) cpsDNA in vivo mirrored xenograft rejection, and correlated with xenograft loss in pig-to-mouse cell transplantation; (d) cpsDNA was significantly reduced when rapamycin was administered; and (e) dynamic cpsDNA was detectable in pig-to-monkey artery patch transplantation. In conclusion, measurement of cpsDNA could prove to be a less invasive, but more specific and sensitive low-cost biomarker enabling monitoring of xenograft rejection and the response to immunosuppressive therapy.

Predictive biomarkers for graft rejection in pig-to-non-human primate corneal xenotransplantation.

We investigated the predictive biomarkers for graft rejection in pig-to-non-human primate (NHP) full-thickness corneal xenotransplantation (n = 34). The graft score (0-12) was calculated based on opacity, edema, and vascularization. Scores ≥ 6 were defined as rejection. NHPs were divided into two groups: (a) graft rejection within 6 months; and (b) graft survival until 6 months. In the evaluation of 2-week biomarkers, none of the NHPs showed rejection within 2 weeks and the 34 NHPs were divided into two groups: (a) entire rejection group (n = 16); and (b) survival group (n = 18). In the evaluation of 4-week biomarkers, four NHPs showing rejection within 4 weeks were excluded and the remaining 30 NHPs were divided into two groups: (a) late rejection group (n = 12); and (b) survival group (n = 18). Analysis of biomarker candidates included T/B-cell subsets, levels of anti-αGal IgG/M, donor-specific IgG/M from blood, and C3a from plasma and aqueous humor (AH). CD8+ IFNγ+ cells at week 2 and AH C3a at week 4 were significantly elevated in the rejection group. Receiver operating characteristic areas under the curve was highest for AH C3a (0.847) followed by CD8+ IFNγ+ cells (both the concentration and percentage: 0.715), indicating excellent or acceptable discrimination ability, which suggests that CD8+ IFNγ+ cells at week 2 and AH C3a at week 4 are reliable biomarkers for predicting rejection in pig-to-NHP corneal xenotransplantation.

What makes D-dimer assays suspicious-heterophilic antibodies?

BACKGROUND: Heterophilic antibodies are still an important source of interference in immunoassays, but reports of interference with D-dimers are rare. Are D-dimer level abnormalities, found in the clinic, caused by heterophilic antibodies as well, or are other mechanisms involved? We will elaborate on this issue through two different examples in this article. METHODS: Serum from two patients with significantly elevated levels of D-dimers were measured and compared by different methods, diluted, and dealt with heterophilic antibody blockers. At the same time, to retrieve the interference, we focused on the cause of D-dimer false positives and made a systematic review of the literature. RESULTS: The D-dimer values were normal (0.49 and 0.15 µg/mL) detected with different testing method and decreased after addition of heterophilic antibody blocking reagent. According to literature data, there were 66.7% (4/6) references showed the interference were heterophilic antibody. CONCLUSIONS: The influence of heterophilic antibodies on the measurement of D-dimers remains a big challenge. Different measuring instruments and methods may have significant differences in the measurement of D-dimers. By using a combination of instrumental methods for measuring, incorporating heterophilic antibody blockers, and combining with clinical performance and imaging data, most of the interference can be eliminated.

Association of open field behavior with blood and semen characteristics in roosters: an alternative animal model / Asociación entre la conducta de gallos en campo abierto con sus características de sangre y de semen: un modelo animal alternativo

Introduction. The paucity of literature is addressed regarding the correlation between open field as an individual behavioral trait on reproductive capacity in animals. Materials and methods. To address this, Nine-month-old indigenous roosters were housed in individual cages. Each animal was observed twice a week for ten minutes before feeding in an open field apparatus for two weeks (7:00-12:00PM). Results. Interestingly, it was found that rooster's semen characteristics were correlated with their open field behavior. On the other hand, plasma glucose level as a blood attribute was more correlated with semen characteristics. The open field monitoring also revealed that the roosters with the lowest delay to their first pace had the highest sperm forward motility and lower sperm abnormality. The heterophil to lymphocyte ratio (H:L) was found to be low when pace bout and pace numbers were 20 and 35, respectively. The negative correlation between H:L ratio and semen characteristics (live sperm percentage, sperm concentration, and membrane integrity) may be an indication of poor reproductive performance in fearful roosters with higher H:L ratio. Conclusions. The data suggested a relationship between open field behavior indices and some reproductive parameters in roosters. The results might be applicable for selection of more reproductive animals. Hence, the rooster may also be useful model for similar studies in other species

Introducción. Existe poca literatura que analice la relación entre el comportamiento individual en animales en campo abierto y su capacidad reproductiva. Material y métodos. Para cubrir esta laguna se trabajó con gallos de 9 meses de edad que fueron encerrados en jaulas individuales. Cada animal se observó 2 veces a la semana, durante 10min, en una zona de campo abierto durante 2 semanas, antes de la alimentación (07:00-12:00 p.m.). Resultados. Curiosamente, se encontró que la característica del semen de los gallos se correlacionaba con su comportamiento en campo abierto. Por otro lado, el nivel de glucosa en plasma, como un atributo de la sangre, se correlacionaba con las características del semen. La monitorización en campo abierto también reveló que los gallos con la menor demora en su primer canto tuvieron la mayor motilidad de los espermatozoides y la menor alteración del esperma. El nivel más bajo de la relación entre heterófilos y linfocitos (H: L) fue encontrado cuando los números de cantos y el ritmo eran 20 y 35, respectivamente. La correlación negativa entre el índice H:L y las características del semen (porcentaje de espermatozoides vivos, concentración de espermatozoides, e integridad de la membrana) puede ser un indicio de mal desempeño reproductivo de gallos con mayor índice H:L. Conclusiones. Los datos indican que existe relación entre los índices de comportamiento en campo abierto y algunos parámetros reproductivos de los gallos. Los resultados podrían ser aplicables para la selección de los animales más reproductivos. El gallo también puede ser útil como un modelo para estudios similares en otras especies

Heterophilic interference in specimens yielding false-reactive results on the Abbott 4th generation ARCHITECT HIV Ag/Ab Combo assay.

BACKGROUND: False-reactivity in HIV-negative specimens has been detected in HIV fourth-generation antigen/antibody or 'combo' assays which are able to detect both anti-HIV-1/HIV-2 antibodies and HIV-1 antigen. OBJECTIVES: We sought to characterize these specimens and determine the effect of heterophilic interference. STUDY DESIGN: Specimens previously testing as false-reactive on the Abbott ARCHITECT HIV Ag/Ab combo assay and re-tested on a different (Siemens ADVIA Centaur HIV Ag/Ab) assay. A subset of these specimens were also pre-treated with heterophilic blocking agents and re-tested on the Abbott assay. RESULTS: Here we report that 95% (252/264) of clinical specimens that were repeatedly reactive on the Abbott ARCHITECT HIV Ag/Ab combo assay (S/Co range, 0.94-678) were negative when re-tested on a different fourth generation HIV combo assay (Siemens ADVIA Centaur HIV Ag/Ab). All 264 samples were subsequently confirmed to be HIV negative. On a small subset (57) of specimens with available volume, pre-treatment with two different reagents (HBT; Heterophilic Blocking Tube, NABT; Non-Specific Blocking Tube) designed to block heterophilic antibody interference either eliminated (HBT) or reduced (NABT) the false reactivity when re-tested on the ARCHITECT HIV Ag/Ab combo assay. CONCLUSIONS: Our results suggest that the Abbott ARCHITECT HIV Ag/Ab combo assay can be prone to heterophilic antibody interference.

Xenoreactive antibodies and latent fibrin formation in VAD and cardiac transplant recipients can confound the detection and measurement of anti-AT1R antibodies.

Autoantibodies to the angiotensin II type 1 receptor (AT1R) are thought to be important in antibody-mediated rejection (AMR), especially in the absence of anti-HLA antibodies. We used a variety of methods to examine the specificity of a commercially available kit designed to quantitate anti-AT1R antibodies. We found that fibrin formation in serum samples from patients awaiting cardiac transplantation with ventricular assist devices (VADs) can produce falsely elevated anti-AT1R values. In addition, absorption studies with a variety of cell lines with or without expression of human AT1R, and those that express xenoantigens, suggest that many of the antibodies detected in the AT1R test system are heterophilic and have reactivity to xenoantigens. Furthermore, we provide data that show that reactivity to the sialic acid Neu5Gc is a common finding among samples that are highest in anti-AT1R levels. We conclude that a common laboratory method for quantitation of anti-AT1R antibodies is nonspecific and overestimates the frequency of true positives. A reevaluation of the role that anti-AT1R antibodies play in allograft function and patient outcomes is warranted.

Peripheral blood detection of systemic graft-specific xeno-antibodies following transplantation of human neural progenitor cells into the porcine spinal cord.

Extensive pre-clinical and clinical studies have searched for therapeutic efficacy of cell-based therapeutics in diseases of the Central Nervous System (CNS) with no other viable options. Allogeneic cells represent the primary source of these therapies and immunosuppressive regimens have been empirically employed based on experience with solid organ transplantation, attempting to avoid immune mediated graft rejection. In this study, we aimed to 1) characterize the host immune response to stem cells transplanted into the CNS and 2) develop a non-invasive method for detecting immune response to transplanted cell grafts. Human neural progenitor cells were transplanted into the spinal cord of 10 Göttingen minipigs, of which 5 received no immunosuppression and 5 received Tacrolimus. Peripheral blood samples were collected longitudinally for flow cytometry cross match studies. Necropsy was performed at day 21 and spinal cord tissue analysis. We observed a transient increase in xeno-reactive antibodies was detected on post-operative day 7 and 14 in pigs that did not receive immunosuppression. This response was not detected in pigs that received Tacrolimus immunosuppression. No difference in graft survival was observed between the groups. Infiltration of numerous immune mediators including granulocytes, T lymphocytes, and activated microglia, and complement deposition were detected. In summary, a systemic immunologic response to stem cell grafts was detected for two weeks after transplantation using peripheral blood. This could be used as a non-invasive biomarker by investigators for detection of immunologic rejection. However, the absence of a detectable response in peripheral blood does not rule out a parenchymal immune response.