1.Screening for Intermediate and Severe Forms of Thalassaemia in Discarded Red Blood Cells: Optimization and Feasibility
Elizabeth George ; Mei I Lai ; Lai Kuan Teh ; Rajesh Ramasamy ; Ern Huei Goh ; Kamalan Asokan ; J A M A Tan ; Maithili Vasudevan ; Sharon Low
The Medical Journal of Malaysia 2011;66(5):429-434
Detection and quantification of Hb subtypes of human blood
is integral to presumptive identification of thalassaemias. It has been used in neonatal screening of thalassaemia and Hb variants. The use of discarded red blood cells following processing of the cord blood for stem cells provides readily available diagnostic material for thalassaemia screening. In this study, we determined the range of Hb subtypes in 195 consecutive cord blood samples collected for cord blood banking. The `cord blood samples’ analysed were those of the remaining red blood cells after the cord blood was processed for stem cell storage. Quantification of Hb subtypes by high performance liquid chromatography (HPLC) was done on BioRad Variant II Hb testing system. Only 73 (36.5%) of the samples could be analyzed neat without dilution. With a 1:300 dilution with wash solution the acceptable area as recommended by the manufacturer for reading of a C-gram within the 1 to 3 million ranges were achieved in all. Eighteen (9%) 12 showed classical Hb Barts (γ4) prerun peaks were confirmed by Sebia Hydrasys automated Hb gel electrophoresis and quantified by Sebia Capillarys 2 capillary electrophoresis. Only 1 (0.5%) was presumptively identified with HbH disease. Due to the
limited number of samples no beta-thalassaemia major, Hb E
beta-thalassaemia and Hb Barts hydrops fetalis were found.
The HPLC assay was possible at a cost US$ 5 per sample and
a turnover time of 10 samples per hour without technical
difficulties. This study reports an effective and valuable
protocol for thalassaemia screening in red blood cells which would otherwise be discarded during cord blood processing. Cord blood with severe and intermediate forms of thalassaemia can be preselected and not stored.
2.Purification and identification of novel cytotoxic oligopeptides from soft coral Sarcophyton glaucum.
Yixian QUAH ; Nor Ismaliza MOHD ISMAIL ; Jillian Lean Sim OOI ; Yang Amri AFFENDI ; Fazilah ABD MANAN ; Lai-Kuan TEH ; Fai-Chu WONG ; Tsun-Thai CHAI
Journal of Zhejiang University. Science. B 2019;20(1):59-70
Globally, peptide-based anticancer therapies have drawn much attention. Marine organisms are a reservoir of anticancer peptides that await discovery. In this study, we aimed to identify cytotoxic oligopeptides from Sarcophyton glaucum. Peptides were purified from among the S. glaucum hydrolysates produced by alcalase, chymotrypsin, papain, and trypsin, guided by a methylthiazolyldiphenyl-tetrazolium bromide (MTT) assay on the human cervical cancer (HeLa) cell line for cytotoxicity evaluation. Purification techniques adopted were membrane ultrafiltration, gel filtration chromatography, solid phase extraction (SPE), and reversed-phase high-performance liquid chromatography (RP-HPLC). Purified peptides were identified by de novo peptide sequencing. From papain hydrolysate, three peptide sequences were identified: AGAPGG, AERQ, and RDTQ (428.45, 502.53, and 518.53 Da, respectively). Peptides synthesized from these sequences exhibited cytotoxicity on HeLa cells with median effect concentration (EC50) values of 8.6, 4.9, and 5.6 mmol/L, respectively, up to 5.8-fold stronger than the anticancer drug 5-fluorouracil. When tested at their respective EC50, AGAPGG, AERQ, and RDTQ showed only 16%, 25%, and 11% cytotoxicity to non-cancerous Hek293 cells, respectively. In conclusion, AERQ, AGAPGG, and RDTQ are promising candidates for future development as peptide-based anticancer drugs.
Amino Acid Sequence
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Animals
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Anthozoa/chemistry*
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Antineoplastic Agents/pharmacology*
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Chromatography, Gel
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Chromatography, High Pressure Liquid
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Chromatography, Reverse-Phase
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Cytotoxins/pharmacology*
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Drug Discovery
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HEK293 Cells
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HeLa Cells
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Humans
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Hydrolysis
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Marine Toxins/pharmacology*
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Oligopeptides/pharmacology*
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Solid Phase Extraction
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Tandem Mass Spectrometry