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Article | 14-October-2020

Enzyme and DTT treatment of adherent RBCs for antibody identification by a solid phase immunoassay system

Treatment of RBCs with protease enzymes or dithiothreitol (DTT) causes denaturation of several RBC antigens and is regularly used in antibody identification. In this study, we have standardized enzyme and DTT treatment of adherent RBCs in the magnetic-mixed passive hemagglutination assay (M-MPHA) for antibody identification. We have also tried drying these treated RBCs. The optimal enzyme and DTT treatment conditions for intact adherent RBCs were determined, in addition to the optimal condition

Toyohiro Tamai, Toshio Mazda

Immunohematology, Volume 18 , ISSUE 4, 114–119

Article | 17-February-2021

Blood component administration to multiple myeloma patients treated with daratumumab: suggesting a novel approach with use of 0.1 M dithiothreitol

, and now guidelines have been laid down by AABB and the British Society for Haematology (BSH) for handling pre-transfusion testing on samples from patients on DARA.4 Treating RBCs with dithiothreitol (DTT) eliminates the DARA interference. DTT denatures the cell surface CD38 by disrupting the disulphide bonds, thus preventing interaction of DARA at the RBC surface and allowing for safe transfusion of patients on DARA.5,6 DTT treatment of RBCs also denatures some of the RBC antigens, such as MER2

P. Pandey, D. Setya, E. Kaul, S. Ranjan, M.K. Singh, A. Shankar

Immunohematology, Volume 36 , ISSUE 4, 157–165

Article | 16-October-2019

Dithiothreitol treatment of red blood cells

Principle Dithiothreitol (DTT) is a reducing agent capable of irreversibly cleaving accessible disulfide bonds when the solution pH is >7. DTT treatment of red blood cells (RBCs) will modify the tertiary structure of protein-based erythrocyte membrane antigens if their confirmation depends on disulfide bonds.1 Antigens of the following blood group systems are destroyed or weakened by 0.2 M DTT treatment: KEL, IN, JMH, YT, LU, MER2, KN, DO, CROM, and LW.2,3 Antibodies directed at antigens in

C.B. Bub

Immunohematology, Volume 33 , ISSUE 4, 170–172

Report | 09-October-2019

Stability guidelines for dithiothreitol-treated red blood cell reagents used for antibody detection methods in patients treated with daratumumab

Daratumumab (DARA), a drug used to treat patients with multiple myeloma, causes interference in pre-transfusion testing. Samples from patients receiving DARA exhibit panreactivity in antibody detection and identification tests with red blood cells (RBCs). Many hospitals are sending these samples to reference laboratories. Dithiothreitol (DTT), a sulfhydryl chemical treatment of RBCs, negates this reactivity. This study investigated the stability of the antigens on DTT-treated RBCs to determine

Wendy L. Disbro

Immunohematology, Volume 33 , ISSUE 3, 105–109

Article | 10-April-2021

Comparative evaluation of the conventional tube test and column agglutination technology for ABO antibody titration in healthy individuals: a report from India

reciprocal to the highest sample dilution showing 1+ agglutination. Dithiothreitol (DTT) is used for the inactivation of IgM antibodies and, therefore, its routine use in immunohematology laboratories is recommended.5 Although the column agglutination technology (CAT) has been described as superior in objectivity and reproducibility when compared with CTT because of the possible overestimation of the antibody titer when using CAT, it has, up to now, only been recommended for antibody identification and

S.S. Datta, S. Basu, M. Reddy, K. Gupta, S. Sinha

Immunohematology, Volume 37 , ISSUE 1, 25–32

Article | 15-February-2021

ZZAP treatment of red blood cells

Principle ZZAP is a mixture of a sulfhydryl reagent (dithiothreitol [DTT]) and a proteolytic enzyme (papain or ficin). It was first described by Branch and Petz in their 1982 article in the American Journal of Clinical Pathology.1 ZZAP is used to dissociate IgG and complement from red blood cells (RBCs), an action that neither reagent can achieve alone. According to Branch and Petz, it is believed that ZZAP “reduces interchain disulfide linkages, increasing exposure of the IgG polypeptides to

S.I. Marckwardt

Immunohematology, Volume 35 , ISSUE 1, 9–10

Original Paper | 09-October-2019

Use of standard laboratory methods to obviate routine dithiothreitol treatment of blood samples with daratumumab interference

, dithiothreitol (DTT) treatment of the patient’s RBCs should be performed. The medical charts of four patients treated with daratumumab were reviewed. The individual number of doses ranged from 1 to 14; patient age ranged from 55 to 78 years; two men and two women were included in the review. Type and screen data were obtained from samples collected over 33 encounters with a range of 1 to 13 encounters per patient. All samples were tested initially by automated solid-phase testing. Any reactivity with

Nicholas J. Lintel, Debra K. Brown, Diane T. Schafer, Farai M. Tsimba-Chitsva, Scott A. Koepsell, Sara M. Shunkwiler

Immunohematology, Volume 33 , ISSUE 1, 22–26

Article | 29-December-2020

Inactivation of Holley (Hy) and Gregory (Gy) antigens by dithiothreitol (DTT)

Robert J. Eckrich

Immunohematology, Volume 4 , ISSUE 1, 12–13

Report | 01-December-2019

The production, serologic evaluation, and epitope mapping of ten murine monoclonal Dombrock antibodies

recognized epitopes that were resistant to treatment with ficin, pronase, α-chymotrypsin, and neuraminidase, but sensitive to trypsin and 0.2 M dithiothreitol (DTT). Five have anti-Dob-like specificity. The epitope recognized by MIMA-52 was neuraminidase sensitive, and MIMA-127 epitope recognized a DTT-resistant, linear epitope 90QKNYFRMWQK99 of the Dombrock polypeptide. MIMA-127 was the only one of the ten Dombrock MoAbs mapped to a specific sequence of the Dombrock glycoprotein; the other nine

Magdalena Grodecka, Kazimiera Wasniowska, Gregory Halverson, Karina Yazdanbakhsh, Marion E. Reid, Elwira Lisowska

Immunohematology, Volume 28 , ISSUE 4, 124–129

Article | 31-December-2020

The Deleterious Effects of Dithiothreitol (DTT) on Red Blood Cell LW Antigens

Ella M. Toy

Immunohematology, Volume 3 , ISSUE 3, 33–35

Article | 16-October-2019

Sulfhydryl treatment of serum or plasma for the reduction of IgM antibodies

Dithiothreitol (DTT) and 2-mercaptoethanol (2-ME) are sulfhydryl compounds that can be used to treat serum or plasma to denature IgM antibody reactivity. By using sulfhydryl agents, IgG and IgM antibodies can be separated, the relative amount of IgM and IgG antibodies can be determined, and the risk of hemolytic disease of the fetus and newborn can be assessed.

Lorraine N. Blagg

Immunohematology, Volume 34 , ISSUE 4, 135–139

Report | 14-March-2020

Absence of hemolytic disease of fetus and newborn despite maternal high-titer IgG anti-Ku

elution of RBC antibodies, and antigen typing. A gravida 3, para 3 (G3P3) woman was first evaluated in 2006 and was found to have an IgG RBC antibody that reacted against all panel RBCs in the anti-human globulin phase. A panel of RBCs treated with DTT did not react with the antibody.  The antibody failed to react with one example of K0 RBCs. The patient’s RBCs typed negative for the following Kell blood group antigens: KEL1, KEL2, KEL3, KEL4, KEL6, KEL7, KEL11, KEL13, and KEL18. These

Ram M. Kakaiya, Angelica Whaley, Christine Howard-Menk, Jigna Rami, Mona Papari, Sally Campbell-Lee, Zbigniew Malecki

Immunohematology, Volume 26 , ISSUE 3, 119–122

Article | 03-November-2020

The gel test: use in the identification of unexpected antibodies to blood group antigens

The IgG GEL test was compared with the LISS tube test (Löw and Messeter’s low-ionic-strength saline) for antibody identification. The suitability of red blood cells (RBCs) pretreated with ficin, dithiothreitol (DTT), or chloroquine diphosphate (CDP) also was assessed for use in the GEL test. In addition, time-in-motion studies were performed comparing GEL (12 panels per batch) with polyethylene glycol (PEG) tube tests (3 panels per batch). In 57 antibody identification studies, there

W. John Judd, E.Ann Steiner, Pamela C. Knaf, Colleen Masters

Immunohematology, Volume 14 , ISSUE 2, 59–62

Article | 30-November-2020

First example of Rh:-32,-46 red cell phenotype

The red cells of a white male blood donor typed as Rh:-1, -2, -3, w4, w5, 6, -17, w19, -31, -32, -34, and -46. Although the donor has no history of transfusion, his serum contains an alloantibody that is weakly reactive with most red blood cells (RBCs) tested. Only Rhnull and D-- RBCs are nonreactive. Reactivity is enhanced with ficin- or papain-treated RBCs and is unaffected by AET or DTT treatment of the RBCs. Previously described Rh:-46 RBCs have been of deletion types D--, D•&bull

Jill Storry, Michael Gorman, Nancy I. Maddox, Ella Toy, Peter D. Issitt, Delores M. Mallory

Immunohematology, Volume 10 , ISSUE 4, 130–133

Article | 30-November-2020

An example of anti-LWa in a 10-month-old infant

Blood samples from a 10-month-old male infant requiring transfusion were found to contain an allomtibody reacting at 37°C in saline, by indirect antiglobulin test (IAT), and with a manual polybrene technique. Preliminary results suggested anti-D and another weaker reacting antibody, but the patient had been previously transfused with only D- blood. His serum reacted more weakly by IAT against red cells treated with 0.2M dithiothreitol (DTT), and one D+, LW(a-) sample was nonreactive. The

Alan Devenish

Immunohematology, Volume 10 , ISSUE 4, 127–129

Case report | 12-March-2020

Role for serial prenatal anti-Vel quantitative serologic monitoring with 2-ME serum treatment during pregnancy: case report

HDN.  Placed in the context of other similar reports, this case demonstrates the importance of separately reporting the IgG fraction (after either DTT treatment or 2-ME treatment) from the untreated (IgM and IgG) fraction and the importance of correlating the treated serum titer with potential clinical significance.

Walter J. Linz, Judith T. Fueger, Steven Allen, Susan T. Johnson

Immunohematology, Volume 26 , ISSUE 1, 8–10

original-paper | 03-September-2019

Purification, Characterization and Inhibition of Alanine Racemase from a Pathogenic Strain of Streptococcus iniae

and different concentrations of hydroxylamine (0.1, 1, and 10 mM) were added to the reaction mixture without the substrate, dialyzed in phosphate buffered saline for 40 min, and its activity was determined without the addition of PLP. The effect of Dithiothreitol (DTT) on the activity of SiAlr was also determined by incubating the enzyme in different concentrations of DTT (1 and 3 mM) for 30 minutes and the relative activity was measured. To confirm SiAlr is a PLP-dependent enzyme, the purified

MURTALA MUHAMMAD, YANGYANG LI, SIYU GONG, YANMIN SHI, JIANSONG JU, BAOHUA ZHAO, DONG LIU

Polish Journal of Microbiology, Volume 68 , ISSUE 3, 331–341

Article | 01-April-2020

An alloantibody to a highprevalence MNS antigen in a person with a GP.JL/Mk phenotype

detected an antigen resistant to treatment by papain, trypsin,α-chymotrypsin, or DTT. The antibody was strongly reactive by the IAT with all RBCs tested except those having the MkMk, GP.Hil/GP.Hil, or GP.JL/GP.JL phenotypes. The patient’s RBCs typed M+N–S+/–s–U+, En(a+/–), Hut–, Mi(a–), Mur–, Vw–, Wr(a–b–), and were TSEN+, MINY+. Reactivity with Glycine soja suggested that her RBCs had a decreased level of sialic acid

John Ratliff, Susan Veneman, Joan Ward, Christine Lomas-Francis, Kim Hue-Roye, Randall W. Velliquette, Laima Sausais, Twilla Maldonado, Janet Miyamoto, Yolanda Martin, David Slater, Marion E. Reid

Immunohematology, Volume 23 , ISSUE 4, 146–149

Article | 14-October-2020

PEG adsorption of autoantibodies causes loss of concomitant alloantibody

Use of polyethylene glycol (PEG) to promote adsorption of autoantibodies is reported to give good recovery of concomitant alloantibodies. In initial experiments, PEG and ZZAP (Ficin and DTT) adsorption procedures were compared for removal of autoantibody and recovery of alloantibody. Postadsorption studies (n = 11) were performed and hemagglutination scores compared. In subsequent studies, equal volumes of alloantibody containing sera, PEG, and antigen-negative red blood cells (RBCs) were used

W. John Judd, Louann Dake

Immunohematology, Volume 17 , ISSUE 3, 82–85

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