DP Typing Research Articles

Patient-derived organoids of pancreatic ductal adenocarcinoma for subtype determination and clinical outcome prediction

BackgroundRecently, two molecular subtypes of pancreatic ductal adenocarcinoma (PDAC) have been proposed: the “Classical” and “Basal-like” subtypes, with the former showing better clinical outcomes than the latter. However, the “molecular” classification has not been applied in real-world clinical practice. This study aimed to establish patient-derived organoids (PDOs) for PDAC and evaluate their application in subtype classification and clinical outcome prediction.MethodsWe utilized tumor samples acquired through endoscopic ultrasound-guided fine-needle biopsy and established a PDO library for subsequent use in morphological assessments, RNA-seq analyses, and in vitro drug response assays. We also conducted a prospective clinical study to evaluate whether analysis using PDOs can predict treatment response and prognosis.ResultsPDOs of PDAC were established at a high efficiency (> 70%) with at least 100,000 live cells. Morphologically, PDOs were classified as gland-like structures (GL type) and densely proliferating inside (DP type) less than 2 weeks after tissue sampling. RNA-seq analysis revealed that the “morphological” subtype (GL vs. DP) corresponded to the “molecular” subtype (“Classical” vs. “Basal-like”). The “morphological” classification predicted the clinical treatment response and prognosis; the median overall survival of patients with GL type was significantly longer than that with DP type (P < 0.005). The GL type showed a better response to gemcitabine than the DP type in vitro, whereas the drug response of the DP type was improved by the combination of ERK inhibitor and chloroquine.ConclusionsPDAC PDOs help in subtype determination and clinical outcome prediction, thereby facilitating the bench-to-bedside precision medicine for PDAC.

Bifurcations, Exact Peakon, Periodic Peakons and Solitary Wave Solutions of Generalized Camassa–Holm–Degasperis–Procosi Type Equation

For the generalized Camassa–Holm–Degasperis–Procosi (CH–DP) type equation, by using the techniques from dynamical systems and singular traveling wave theory developed by Li and Chen [2007] to analyze its corresponding traveling wave systems, which depend on four parameters, it is found that under different parameter conditions its bifurcation portraits exhibit all possible exact explicit bounded solutions, such as solitary wave solutions, periodic wave solutions, peakon as well as periodic peakons. A total of 30 explicit exact parametric representations of the traveling wave system of the CH–DP type equation are presented.

Knowing and Believing Things: What DP-Complementation Can Tell us about the Meaning and Composition of (Factive) Attitudes

Abstract In the Hintikkan tradition, attitude verbs are viewed as relations between individuals and propositions. Previous work on know and believe with Content DPs like the rumour has tended to treat know CP vs. know DP as polysemy. In this paper, I show that polysemy runs into conceptual and empirical problems, and propose instead a new decompositional approach to know-verbs, which avoids polysemy; linking both know DP and know CP to the same lexical root, which describes, broadly speaking, acquaintance. This analysis thus provides an explicit and compositional morpho-semantic link between know DP and know CP that accounts for the interpretation of DPs as objects of acquaintance, and further captures the idea that knowledge, and factivity more broadly, is tied to acquaintance with a situation, the res ( Kratzer 2002, a.o.). Based on detailed examination of the morpho-syntax and interpretation of DP and CP complements of believe, I further show that DPs can either combine with believe in the same fashion as CPs, as a direct object (saturating a propositional argument slot, as in Uegaki 2016), or as an indirect object, via a type of attitudinal applicative (proposed here). The former option is defined for Content DPs and the latter for agentive DPs, so-called Source DPs. Together, these proposals account for the observation that the interpretation of believe DP sentences varies depending on the type of DP (believe the rumour vs. believe the referee), whereas for know-verbs, both types of DPs are interpreted as objects of acquaintance. At the core of the current proposal is the idea that verbs like know and believe differ fundamentally at the level of argument structure and internal morpho-semantic composition, and thus combine with DPs via different routes; contrary to uniform approaches to know and believe. Whereas believe-verbs describe relations to intensional content, and require external licensing mechanisms to combine with DPs, know-verbs describe complex relations, fundamentally anchored in the attitude holder’s acquaintance with (abstract or concrete) individuals in the world, and thus make reference to individuals as part of their argument structure. The current proposal also builds on and adds to previous insights about connections between factivity, DP-complementation, and question-embedding.

Unrelated donor α/β T cell– and B cell–depleted HSCT for the treatment of pediatric acute leukemia

Unrelated donor (URD) hematopoietic stem cell transplant (HSCT) is associated with an increased risk of severe graft-versus-host disease (GVHD). TCRαβ/CD19 depletion may reduce this risk, whereas maintaining graft-versus-leukemia. Outcome data with TCRαβ/CD19 depletion generally describe haploidentical donors, with relatively few URDs. We hypothesized that TCRαβ/CD19-depletion would attenuate the risks of GVHD and relapse for URD HSCT. Sixty pediatric and young adult (YA) patients with hematologic malignancies who lacked a matched-related donor were enrolled at 2 large pediatric transplantation centers between October 2014 and September 2019. All patients with acute leukemia had minimal residual disease testing, and DP typing was available for 77%. All patients received myeloablative total body irradiation– or busulfan-based conditioning with no posttransplant immune suppression. Engraftment occurred in 98%. Four-year overall survival was 69% (95% confidence interval [CI], 52%-81%), and leukemia-free survival was 64% (95% CI, 48%-76%), with no difference between lymphoid and myeloid malignancies (P = .6297 and P = .5441, respectively). One patient (1.7%) experienced primary graft failure. Relapse occurred in 11 patients (3-year cumulative incidence, 21%; 95% CI, 11-34), and 8 patients (cumulative incidence, 15%; 95% CI, 6.7-26) experienced nonrelapse mortality. Grade III to IV acute GVHD was seen in 8 patients (13%), and 14 patients (26%) developed chronic GVHD, of which 6 (11%) had extensive disease. Nonpermissive DP mismatch was associated with higher likelihood of acute GVHD (odds ratio, 16.50; 95% CI, 1.67-163.42; P = .0166) but not with the development of chronic GVHD. URD TCRαβ/CD19-depleted peripheral HSCT is a safe and effective approach to transplantation for children/YAs with leukemia. This trial was registered at www.clinicaltrials.gov as #NCT02323867.

Definiteness in Laki: Its interaction with demonstratives and number

AbstractThis article investigates definiteness and its interactions with demonstratives and number in Laki (Northwestern Iranian). By the examination of demonstratives and building upon previous proposals, I argue for two types of definite DPs in Laki, namely anaphoric and deictic. I show that the patterns of definite and number marking are sensitive to the type of the DP. In particular, I argue that double definiteness, resulting from an Agree relation between D and N, and head movement of Num to D both are obtained only in anaphoric definite DPs for feature-checking requirements. Overall, this study highlights the contributions of anaphoricity to the DP internal structure. The present proposal can account for similar phenomena in other Iranian languages (i.e., Sorani and Kermanshahi Kurdish). The divergence of Laki definiteness from similar attested patterns (i.e., Scandinavian double definiteness) contributes to our cross-linguistic understanding of definiteness and its interactions with other nominal elements.

Substituting imputation of HLA antigens for high-resolution HLA typing: Evaluation of a multiethnic population and implications for clinical decision making in transplantation.

Molecular mismatch analysis for assessment of histocompatibility in transplantation requires high-resolution HLA typing. Algorithms to "guesstimate" high-resolution from low-resolution typing exist, but their accuracy remains unknown. We converted high-resolution, sequence-based, HLA typing of 310 subjects from an ethnically heterogeneous population to low-resolution equivalents and tested the ability of the NMDP HaploStats and HLA Matchmaker programs to impute/reproduce the measured high-resolution HLA type, using the more common "winner-takes-all" approach. Only 35.6% of the HaploStats imputed HLA-A, -B, -C, -DRB1, and -DQB1 haplotypes had no mistakes, and the accuracy was significantly lower for non-Caucasians (29.1%) compared to Caucasians (45.2%) (odds ratio [OR], 0.5; 95% confidence interval [CI], 0.3-0.8; P=.004). HLA Matchmaker was not able to provide high-resolution haplotypes for 45.2% of Caucasian subjects and 63.5% of non-Caucasian subjects (P=.002). Of those with an imputed result, only 10.3% of Caucasians and 4.8% of non-Caucasians had accurate 10-allele high-resolution output. Eplet analysis revealed additional, inaccurate eplets in 37% of individuals, with 22.5% showing at least 2 additional, inaccurate eplets; incorrect eplets were more common among non-Caucasians (OR, 1.8; 95% CI, 1.1-2.9; P=.018). Given this high error rate, caution should be taken before using imputation tools for clinical or research purposes, especially for non-Caucasian individuals.

Assessment by Extended-Coverage Next-Generation Sequencing Typing of DPA1 and DPB1 Mismatches in Siblings Matching at HLA-A, -B, -C, -DRB1, and -DQ Loci.

Allogeneic hematopoietic stem cell transplant from an HLA matched sibling donor is usually the preferable choice. The use of next-generation sequencing (NGS) for HLA typing in clinical practice provides broader coverage and higher resolution of HLA genes. We evaluated the frequency of DPB1 crossing-over events among patients and potential related donors typed with NGS. From July 2016 to January 2018, 593 patients and 2385 siblings were typed. We evaluated sibling matching status in 546 patients, and 44.8% of these patients had siblings that matched at HLA-A, -B, -C, -DRB1, and -DQB1 loci. In 306 patient-HLA matched sibling pairs, we found 6 pairs (1.96%) with 1 DPB1 mismatch, and 5 of these pairs included an additional mismatch in DPA1. No additional mismatches were observed at the low expression loci. Using the T cell epitope algorithm, 4 of these DP mismatches were classified as permissive, 1 as nonpermissive in the host-versus-graft direction, and 1 as nonpermissive in the graft-versus-host direction. The frequency of DPB1 and DPA1 mismatches is low, and their impact in related donor transplants is not well established. Although DP typing in related transplants goes beyond guidelines, it is especially relevant for sensitized patients. NGS-based HLA typing provides full gene coverage, and its use in clinical practice can enable better donor selection.

Lower and Higher Copies in Arabic

This paper examines the phenomenon of copying in Standard Arabic (SA) within the copy theory. Following Boskovic and Nunes (2007), the researcher contends that Arabic utilizes two ways for the pronunciation of heads and copies governed by syntactic and phonological constraints. The highest link can be phonetically realized, or copies are pronounced instead of their heads by PLC without causing the derivation to crash. The paper shows how the copy theory accounts for the differences between perfective and imperative forms on one hand and the imperfective form on the other hand. The copy theory helps account for V-to-T0 movement in imperatives and perfectives in comparison with remaining in lower projections in imperfectives. Further, the research reveals that certain fixed word orders in Arabic can be accounted for in terms of the copy theory. The type of the object DP plays a role in which copy is realized, and hence affects word order. The higher copy of the object DP is pronounced when it is an enclitic object pronoun, while the lower copy is pronounced when the object is an R-expression DP. Addressing the topic of copying from a semantics-syntax interface perspective, the paper explains the bearings of copy operations on syntax..

P050 Unusual DP alleles are being found on a regular basis and may not be “unusual”

An African American deceased donor was typed by Luminex rSSO (One Lambda) as A23, 30; B45, 52; Cw6, 16; DR7, 18; DR 52, 53; DQ 2, 4; DPB1*01:01, 425/522. Follow-up re-typing of the DP locus by SSP showed DRB1*01:01, 354:01/375:01/584:01. The second DP did not share any alleles between the SSP and rSSO strings, making it impossible to call a type for the second allele. Interestingly, another African American donor showed the same unusual strings by rSSO and SSP within a few weeks of the first typing. The occurrence of these unusual and discrepant strings (SSP vs rSSO) in two patients of the same ethnicity suggested that this may not be a rare allele and identification of this allele is needed. The sample results were sent to the manufacturer who reported that DPB1*584:01 was a shared allele with both methods and was the probable allele. However, our Luminex analysis did not show DPB1*584:01 as an ambiguity with either of these samples. NGS was requested to confirm the DP type. This case demonstrates some of the difficulties laboratories face when performing DP typing, especially on deceased donor samples. More and more “unusual” DP typings are being found on a regular basis. We do not yet have a clear understanding of which DP antigens are unusual or rare. Analysis software for a particular lot may not be updated in a timely fashion to include the latest alleles that should be in the list of ambiguities. Confirmation of these unusual antigens is also not possible during the time constraints of performing deceased donor typing. An even greater problem may be that the DP antibody testing is drastically behind in identifying antibodies to an ever-growing number of possibly “common” DP antigens. Even if we were able to confirm the DP in this sample as DPB1*584:01, we would not be able to identify specificity against this particular antigen, using the currently available DP antibody reagents.

P026Timing is everything – Even for DP antibodies

Introduction Donor Specific Antibodies (DSA) can be a contraindication for kidney transplants. The clinical relevance of DSA to DP antigens and correlation with crossmatch (XM) remain unclear. We report the conversion of a negative XM to positive due to development of DSA to DP6, most likely caused by an unreported sensitizing event. Methods A 38 yo female received a deceased donor (DD) kidney transplant (Tx) after negative T/B cell flow XMs using sera drawn 35 and 44 days pre Tx (day −35 and −44). Delta channel shifts (DCS) in these XMs were: T = 0 for both sera; B = 37 and 31 for days −35 and −44. These sera were negative for DSA. We received a serum drawn at admission for Tx (day −1). The patient received blood transfusions within the previous week (day −6). The day-1 serum had high level DSA to DP6 (MFI = 13,723), compared to day −44 (1,321) and day-35 (1,653). By flow XM, the day −1 serum was T negative, B positive (DCS = 228), likely due to the increase in DSA to DP6. In addition to standard immunosuppression, this recipient also received thymoglobulin and basiliximab once the program was notified of high level DSA and positive flow XM. Post Tx monitoring showed an immediate decline in DSA to DP6: post Tx day 6 (MFI = 7,380), day 12 (5,669), day 21 (4,042), day 63 (485). The DSA to DP6 remained low (MFI 400) for five years. However, the recipient had BK viremia and a kidney biopsy five years post Tx showed “arteriolar hyalinosis and arteriosclerosis related to possible CNI toxicity”. The recipient is re-listed in UNET for another kidney Tx, the primary reason for graft failure deemed transplant glomerulopathy. Discussion This case emphasizes the importance of timely antibody testing in relation to XM testing, awareness of sensitizing events, and updated HLA typing. Our lab now employs stricter guidelines to ensure more frequent receipt of patient sera, especially following potential sensitizing events. At the time of this patient’s Tx, DP typing was not performed on DD. After discovering the increase in antibodies at pre Tx day −1, but absence of DSA, we performed DP typing. Finding DP6 as a DD antigen helped explain the change to positive XM, as the level of DSA to DP6 significantly increased following the sensitizing event pre-Tx. This early peri and post Tx antibody response may have contributed to reduced graft survival and transplant glomerulopathy.

Sequence-based HLA-A, B, C, DP, DQ, and DR typing of 159 individuals from the Worcester region of the Western Cape province of South Africa

DNA sequence-based typing at the HLA-A, -B, -C, -DPA1, -DPB1, -DQA1, -DQB1, -DRB1, and -DRB3/4/5 loci was performed on samples provided by 159 individuals from the Worcester region of the Western Cape province of South Africa. The purpose of the study was to characterize allele frequencies in the local population, to support studies of T cell immunity against pathogens, including Mycobacterium tuberculosis. There are no detectable deviations from Hardy Weinberg proportions for the HLA-A, -B, -C, -DPA1, -DPB1, -DQA1, and -DRB1 loci. A minor deviation was detected at the HLA-DQB1 locus due to an excess of homozygotes. The genotype data are available in the Allele Frequencies Net Database under identifier 3425.

The dp type π-bond and chiral charge density waves in 1T-TiSe2.

Based on the atomic electronic configuration and Ti-Se coordination, a valence bond model for the layered transition metal dichalcogenide (TMDC) 1T-TiSe2 is proposed. 1T-TiSe2 is viewed as being composed of edge-sharing TiSe4-plaquettes as TiSe2-ribbon chains in each layer via a directional valence shell electron distribution as chemical bonds, in contrast to the conventional layer view of face-sharing TiSe6-octahedra. The four valence electrons per Ti in the hybridized dsp2-orbitals of square coordination form σ-bonds with the four nearest neighbor Se atoms in the chain. The electrons in the lone pair of the Se-4pz orbital are proposed to form a dp type π-bond via side-to-side orbital overlap with the empty Ti-3dxz/3dyz orbitals within each chain, which is positively supported by quantum chemistry calculations. A study of electron energy loss spectroscopy (EELS) with transmission electron microscopy (TEM) for 1T-TiSe2 is presented to show an energy loss near ∼7 and ∼20 eV, which confirms the existence of collective plasmon oscillations with the predicted effective electron numbers for the π- and (π + σ)-bond electrons, respectively.

P109 Clinical outcome of hematopoietic stem cell transplantation -HSCT in patients with DP-TCE matched donors and various stem cell sources

Aim Attaining an HLA matched HSCT is important in achieving better graft survival. This study focuses on comparing patients with DP-TCE permissive transplants to those with unknown DP-TCE, to understand the impact of DP typing on graft versus host disease (GVHD). The source of stem cells is included in this study to evaluate any impact it may have on clinical outcomes. Methods Patients: We analyzed 110 consecutive patients that underwent HSCT at OU Medical Center from 2014 to 2016. In this cohort, DP typing was available in 15 and DP was unknown in remaining. Clinical observation: Charts were reviewed and GVHD assessment was collected after transplant. Statistic: Fisher exact test. Results Transplants from unrelated donors with 10/10 HLA matched and DP-TCE permissive (N = 10) graft developed GVHD in 60% of cases while transplants from DP-TCE non-permissive (N = 5) graft developed GVHD in 100% of cases. The majority of patients that underwent DP typing were peripheral blood (73%). Prior to DP typing, patients that were 10/10 matched to an unrelated donor showed less GVHD (P value: 0.038) when transplanted with bone marrow (N = 34) versus peripheral blood (N = 32) stem cells. Similar findings were seen in siblings that were 10/10 matched (see table). As expected, cord blood fared better with mismatches, confirming data that mismatched cord blood transplant GVHD was similar to unrelated donor transplants. Conclusions Patients with DP-TCE non-permissive transplant are more likely to develop GVHD compared to patients with DP-TCE permissive transplant. Peripheral blood stem cell transplant patients showed higher levels of GVHD than other cell sources. Matched related donors showed decreased levels of GVHD compared to matched unrelated donors, and cord blood showed lower level of GVHD. DP-TCE testing, however, identified subgroups within the transplanted cohort that fared better if permissive typing was found. The number of cases identified is small, but warrants larger assessment to confirm observations seen. Download : Download high-res image (116KB) Download : Download full-size image

P135 New frequent HLA-DPB1/DPA1 haplotypes in low resolution typing

Aim The linkage between DPB1 and DPA1 was extensively studied in 5,944 European American stem cell donors (Hollenbach et al., 2012). Thirty-three DPB1 alleles and associated DPA1 were reported. In 2014, Voorter et al. reported the special association of DPB1∗105:01 with DPA1∗03:01 in contrast to DPB1∗04:02-DPA1∗01:03 association although both DPB1∗105:01 and DPB1∗04:02 belong to the same G group. Here we retrospectively reviewed 1938 renal recipients with DP typing and report new DPB1/DPA1 associations frequently seen in our cohort. Methods HLA-DPB1 and DPA1 typing was performed by rSSO (One Lambda, Thermo-Fisher). Haplotypes were inferred by Clark’s method published in 1990. Results We identified 11 DPB1 alleles whose association with DPA1 has never been reported previously (Table 1). Five DPB1 alleles, DPB1∗124:01, 126:01, 131:01, 350:01 and 352:01, are not in the 2013 CWD list. Of note, DPB1∗131:01 was detected at high frequency (2.9%) in our cohort. The association of these five DPB1s with DPA1 was the same as the most frequent allele in the same G group, e.g. DPB1∗126:01 and DPB1∗350:01 are both associated with DPA1∗01:03 the same as DPB1∗04:01. Six DPB1 CWD alleles observed in our cohort were not reported in Hollenbach’s study. In 2015, Aaron et al. reported significant variation of DPB1 frequency in four major ethnic groups. In their study, DPB1∗29:01, 39:01, 40:01, and 85:01 were much more frequent in African American than other three ethnic groups. High percentage of African American in our renal recipients (44%) may explain the detection of these alleles in our cohort. Interestingly DPB1∗106:01 was associated with DPA1∗02:01, unlike PB1∗19:01 which is in the same G group however is associated with DPA1∗02:02. Conclusions We reported 11 new DPB1/DPA1 haplotypes frequently seen in DP low resolution typing. Five non-CWD DPB1 alleles were identified frequently. The enrichment of some alleles in our cohort could be attributed to high incidence of African American in our patients. Download : Download high-res image (190KB) Download : Download full-size image

Position and orientation of gallated proanthocyanidins in lipid bilayer membranes: influence of polymerization degree and linkage type

It is well known that the biological activity of gallated proanthocyanidins (PAs) is highly structure-dependent. Polymerization degree (DP) and linkage types affect their biological activity greatly. Positions and orientations of gallated PAs in lipid bilayer reveal their structure-function activity at the molecular level. The present work aimed at determining the locations and orientations of epigallocatechin-3-gallate (EGCG) and its derivatives: A-type and B-type EGCG dimers and trimers in 1-palmitoyl-2-oleoyl phosphatidylcholine (POPC) and 1-palmitoyl-2-oleoyl phosphatidylethanolamine (POPE) lipid bilayer via molecular dynamic (MD) simulations. The results showed that EGCG and its derivatives localized in the lipid bilayer or on the bilayer/water interface. Their penetration depths and orientations depended on both DP and linkage types. The penetration depths decreased with the increase of DP, sequencing to be EGCG > EGCG dimers > EGCG trimers. Spatially stretched A-type PAs could form more hydrogen bonds (H-bonds) with deep oxygen atoms of lipid bilayer and have higher affinity to the lipid bilayer than B-type PAs. Our results will provide an explicit evidence for PAs’ distinct biological activities.

Sequence-based HLA-A, B, C, DP, DQ, and DR typing of 100 Luo infants from the Boro area of Nyanza Province, Kenya

One hundred healthy infants enrolled as controls in a tuberculosis vaccine study in Nyanza Province, Kenya provided anonymized samples for DNA sequence-based typing at the HLA-A, -B, -C, -DPB1, -DQA1, -DQB1, -DRB1, and -DRB3/4/5 loci. The purpose of the study was to characterize allele frequencies in the local population, to support studies of T cell immunity against pathogens, including Mycobacterium tuberculosis. There are no detectable deviations from Hardy Weinberg proportions for the HLA-B, -C, -DRB1, -DPB1, -DQA1 and -DQB1 loci. A minor deviation was detected at the HLA-A locus due to an excess of HLA-A*02:02, 29:02, 30:02, and 68:02 homozygotes. The genotype data are available in the Allele Frequencies Net Database under identifier 3393.

OR52 HLA typing from a deceased donor who received hematopoietic stem cell transplant

Here we present a case of challenging typing for a deceased donor who is also hematopoietic stem cell (HSC) transplant recipient. A local deceased donor (DD) was typed with SSP method from peripheral blood sample as A2; B44, 62; Cw9,16; DR4,7; DQ2,7; DQA1∗02, 03; DPB1∗04:02, 11:01; DPA1∗01, 02. After typing was reported, it was immediately found out DD received HSC transplant from an unrelated donor in 2001 at another center. DD has full engraftment on death so HLA typing based on peripheral blood sample is likely the typing for her HSC donor. Typing for DD before HSC was found to be: A2; B44, 62; CW5, 9; DR4,7; DQ7,9, while her HSC donor is A2; B44, 62; Cw9,16; DR4,7; DQ2,7. DQA1∗ and DP typing were not performed in 2001 and no stored DNA can be found. There are two mismatched antigen on HLA-CW and -DQ. This confirmed that initial typing results for DD in fact were typing for her HSC donor, not typing for the kidneys. One kidney was exported to a patient with anti-HLA DP antibodies at another center. The receiving center asked to determine HLA-DP phenotype for DD. Without other source of somatic DNA, a piece of blood vessel from harvested kidney was used to extra DNA for DPB1∗ SSP typing. The blood vessel was contaminated with blood from her HSC donor. It was found that there are four DPB1 antigens: DPB1∗04:02, 11:01, and DPB1∗01:01, 04:01. HSC donor typing is the typing got from blood sample of DD: DPB1∗04:02, 11:01. So DPB1∗01:01, 0401 were deduced as the presumptive typing for this deceased donor. Fortunately there is no anti-HLA-DP DSA for the designated recipient. In summary, a deceased donor was mistakenly typed based on improper source of specimen. Recipients of HSC transplant rarely become deceased organ donors. However, many patients received HLA-DP mismatched unrelated HSC transplant because usually HLA-DP was not matched. Anti-HLA-DP antibodies are found in lots of patients waiting for kidney transplant. This case highlights the importance of communication with deceased donor team, and proper sample collection to achieve accurate deceased donor typing and excellent virtual cross match.

OR34 Examination of HLA DP inton/exon variation identifies two DPB1 evolutionary groups including intron 2 STR variants that may regulate DPB1 expression levels

Recent studies showed the existence of biologically relevant DPB1 expression variants that associate with two 3′UTR SNPs of DPB1. The knowledge of sequence variation in DPA1 and DPB1 alleles is limited to a few exons. Only a few alleles are fully sequenced resulting in limitations for DP typing by NGS. We developed robust and efficient methods for long range amplification and molecular cloning that allowed us to examine and characterize numerous alleles. Clones were sequenced by NGS and consensus was generated by a novel de novo assembly algorithm. The fragments generated span exon 1–4 and 2–4 of DPA1 and DPB1. We characterized 16 DPA1 and 40 DPB1 alleles defined but their unique sequences in 63 samples. We obtained novel sequence information in 11 DPA1 alleles including 7 extended sequences, 3 novel intron variants and 1 novel exon variant. In DPB1 we extended the coverage in 20 allleles and identified 12 novel intron variants and 2 novel exon variants. In addition to the well known variations in exons, the long DPA1 intron1 (3556–3590 nt) showed 201 SNPs and 17 indels. DPA1 intron 2 (340 nt) and intron 3 (214 nt) are short, not variable in size with low polymorphism (12 and 9 SNP, respectively). In DPB1 intron 3 is conserved in length (547 nt) and presents only 17 SNPs. Intron 2 of DPB1 (3952–4025 nt) has the largest variability with 176 SNPs and 9 indels; in addition this intron has one variable STR region located at 44 nt before the beginning of exon 3 with 4–14 repeats of the tetramer AAGG. Two groups of alleles that have only 4 repeats (short STR) and 8–14 repeats (long STR) were identified. Also, phylogenetic trees define two distinct groups at each, DPA1 and DPB1 loci. The DPB1 clusters are tighter when exon 2 is excluded from the analysis and correlate well with the intron 2 STR short and long variants. We observed that the 3′ UTR variants of DPB1 that correlate with high and low expression, also associate strictly in a biunivocal fashion with the short and long intron 2 STR variants, respectively. Our studies provide useful information for mapping reads in NGS typing. In addition our findings identify potential regulatory elements that may affect gene expression possibly by altering the rate of mRNA splicing. In transplantation, the expression levels of the mismatched allele appear to determine both immunogenicity and outcome.

Microstructural Aspects of Twin-Spot Laser Welding of Dp-Hsla Steel Sheet Joints

Abstract The article presents the possibility of using twin-spot laser welding (i.e. laser welding with focusing a laser beam on two spots) for making overlap joints made of 0.8-1 mm thick HSLA and DP type high strength steel sheets. Joints were made using a Yb:YAG disc laser having a maximum power of 12 kW and a welding head by means of which it was possible to focus a laser beam on two spots, 0.6 mm and 1 mm away. The angle between focuses amounted to 0° or 90°, whereas the power distribution was 50%-50%. With settings as presented above it was possible to obtain high-quality overlap joints. The geometrical parameters of the joints were primarily affected by beams positions (in relation to each other) and, to a lesser degree, by the distance between the focuses. It was possible to obtain a 10% hardness reduction in the fusion zone of the DPHSLA steel joints if the angle between the beams amounted to 90°. The tests also involved microstructural examinations of individual zones of the joints

Comparative structural characterization of 7 commercial galacto-oligosaccharide (GOS) products

Many β-galactosidase enzymes convert lactose into a mixture of galacto-oligosaccharides (GOS) when incubated under the right conditions. Recently, the composition of commercial Vivinal GOS produced by Bacillus circulans β-galactosidase was studied in much detail in another study by van Leeuwen et al. As a spin-off of this study, we used the developed analytical strategy for the evaluation of 6 anonymous commercial GOS products, in comparison with Vivinal GOS. These GOS products were first subjected to HPLC-SEC, calibrated HPAEC-PAD profiling (glucose units in relation to a malto-oligosaccharide ladder), and 1D 1H NMR spectroscopy. For a more detailed analysis and support of the conclusions based on the initial analysis, the GOS products were separated into DP-pure subpools on Bio-Gel P-2 (MALDI-TOF-MS analysis), which were subjected to calibrated HPAEC-PAD profiling and 1H NMR analysis. Unidentified peaks from different GOS products, not present in Vivinal GOS, were isolated for detailed structural characterization. In this way, the differences between the various GOS products in terms of DP distribution and type of glycosidic linkages were established. A total of 13 new GOS structures were characterized, adding structural-reporter-group signals and HPAEC-PAD based glucose unit G.U. values to the analytical toolbox. The newly characterized products enhance the quality of the database with GOS structures up to DP4. The combined data provide a firm basis for the rapid profiling of the GOS products of microbial β-galactosidase enzymes.