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Fulltext Identification of major and minor allergens of black tiger prawn (Penaeus monodon) and king prawn (Penaeus latisulcatus)

Sahabudin S, Misnan R, Yadzir ZH, Mohamad J, Abdullah N, Bakhtiar F, et al.

Malays J Med Sci, 2011 Jul;18(3):27-32.
PMID: 22135598 MyJurnal

BACKGROUND: Prawns and shrimp are a frequent cause of seafood allergy mediated by IgE antibodies. Penaeus monodon and Penaeus latisulcatus, commonly known as black tiger prawn and king prawn, respectively, are among the most frequently consumed prawns in Malaysia. The aim of this study was to identify the IgE-binding proteins of these 2 prawn species.
METHODS: Raw and boiled prawn extracts were prepared and then resolved by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE). IgE-immunoblotting was then performed using sera from patients with positive skin prick tests to the raw prawn extracts.
RESULTS: SDS-PAGE analysis of the raw extracts of both prawn species revealed 23 protein bands; the boiled extracts yielded fewer protein bands. The bands in the range of 40 to 100 kDa were sensitive to heat and therefore were not found in the boiled extracts. Immunoblot of raw extracts of black tiger prawns and king prawns yielded 14 and 11 IgE-binding proteins, respectively, with molecular weights of between 15 and 200 kDa. Proteins at 36, 42, and 49 kDa were detected as the major allergens in both species of prawns. A protein of 75 kDa was also identified as a major allergen in black tiger prawns. Other potential allergens were also observed at various molecular masses.
CONCLUSION: Proteins of 36, 42, and 49 kDa were identified as the major allergens of both species of prawns. The 36 and 42 kDa proteins are hypothesised to be tropomyosin and arginine kinase, respectively. A high molecular weight protein of 75 kDa was found to be an additional major allergen in black tiger prawns.
KEYWORDS: Penaeus; allergens; allergy and clinical immunology; hypersensitivity; immunoblotting; tropomyosin

Matched MeSH terms: Electrophoresis, Polyacrylamide Gel
Inhibition of hepatitis B virus assembly with synthetic peptides derived from the viral surface and core antigens

Tan WS

J Gen Appl Microbiol, 2002 Apr;48(2):103-7.
PMID: 12469306

The long surface antigen (L-HBsAg) of hepatitis B virus (HBV) plays a central role in the production of infectious virions. During HBV morphogenesis, both the PreS and S domains of L-HBsAg form docking sites for the viral nucleocapsids. Thus, a compound that disrupts the interaction between the L-HBsAg and nucleocapsids could serve as a therapeutic agent against the virus based upon inhibition of morphogenesis. Synthetic peptides correspond to the binding sites in L-HBsAg inhibited the association of L-HBsAg with core antigen (HBcAg). A synthetic peptide carrying the epitope for a monoclonal antibody to the PreS1 domain competed weakly with L-HBsAg for HBcAg, but peptides corresponding to a linear sequence at the tip of the nucleocapsid spike did not, showing that the competing peptide does not resemble the tip of the spike.

Matched MeSH terms: Electrophoresis, Polyacrylamide Gel
Immunochemical characterization of edible bird's nest allergens

Goh DL, Chua KY, Chew FT, Liang RC, Seow TK, Ou KL, et al.

J Allergy Clin Immunol, 2001 Jun;107(6):1082-7.
PMID: 11398089

BACKGROUND: We have previously described anaphylaxis induced by edible bird's nest (BN) and demonstrated that this condition is IgE mediated.
OBJECTIVES: This study aimed at describing the immunochemical properties of the BN allergens. Comparative studies between 3 commercially available sources (according to the country of origin) of BN were also made.
METHODS: Crude extracts of commercially available processed BN from Sarawak (Malaysia), Thailand, and Indonesia and fresh unprocessed BN from the caves of Sarawak were obtained by means of aqueous extraction. Specific IgE toward these sources were determined by using fluorescence allergosorbent tests (FASTs). Cross-reactivity studies between the 3 sources of commercially available processed BN were carried out by means of FAST inhibition. Immunochemical characterization by means of IgE immunoblot, periodate treatment, and heat stability studies were carried out on fresh unprocessed BN from Sarawak.
RESULTS: Serum from allergic patients showed differences in IgE binding to the 3 sources of commercially available BN, with the highest levels of specific IgE recorded with the Sarawak source (P

Matched MeSH terms: Electrophoresis, Polyacrylamide Gel
Diagnosis of natural rubber latex allergy: multicenter latex skin testing efficacy study. Multicenter Latex Skin Testing Study Task Force

Hamilton RG, Adkinson NF

J Allergy Clin Immunol, 1998 Sep;102(3):482-90.
PMID: 9768592

BACKGROUND: No characterized diagnostic natural rubber latex skin testing material is licensed for use in the United States.
OBJECTIVE: We have conducted a multicenter clinical skin testing study to document the safety and diagnostic sensitivity and specificity of a candidate Hevea brasiliensis nonammoniated latex (NAL) extract. These data are intended to support the licensing of this reagent for the diagnosis of latex allergy in high-risk populations.
METHODS: Three hundred twenty-four subjects (304 adults and 20 children) were classified by their clinical history as having latex allergy (LA group, 124 adults and 10 children) or having no latex allergy (NLA group, 180 adults and 10 children). All subjects provided blood samples and then received sequential puncture skin tests (PSTs) at 1, 100, or 1000 microg/mL protein with a bifurcated needle and NAL (Greer Laboratories) from Malaysian Hevea brasiliensis (clone 600) sap. A 2-stage glove provocation test was used to clarify latex allergy status of individuals with positive history/negative PST result and negative history/positive PST result mismatches.
RESULTS: Twenty-four subjects (15%) originally designated as having LA on the basis of their initial clinical history were reclassified to the NLA group on the basis of a negative glove provocation test result. Of the 134 subjects with LA, 54 (40%) were highly sensitive to latex, with a positive PST result at 1 microg/mL NAL. The Greer NAL reagent produced a positive PST rate (sensitivity) of 95% and 99% in subjects with LA at 100 microg/mL and 1 mg/mL, respectively. The negative PST rate (specificity) in 190 subjects with a negative history with the NAL extract at 100 microg/mL and 1 mg/mL, was 100% and 96%, respectively. Immediately after the PST, mild systemic reactions (mainly pruritus) were recorded in 16.1 % of the adults in the LA group and 4.4% of the adults in the NLA group. No reactions required treatment with epinephrine. Only mild delayed reactions were observed in 9.6% (LA group) and 2.8% (NLA group) of subjects 24 to 48 hours after PST. Mean wheal and erythema diameters measured in the 10 children in the LA group with spina bifida at 100 microg/mL and 1 mg/mL were similar to those observed in the adults in the LA group, suggesting that children are not at increased risk for systemic reactions compared with adults.
CONCLUSIONS: A suggestive clinical history is necessary but not sufficient for a definitive diagnosis of IgE-dependent latex allergy. These data support the safety and diagnostic efficacy of the Greer NAL, skin test reagent at 100 micro/mL and 1 mg/mL for confirmatory PSTs.

Matched MeSH terms: Electrophoresis, Polyacrylamide Gel
The rubber elongation factor of rubber trees (Hevea brasiliensis) is the major allergen in latex

Czuppon AB, Chen Z, Rennert S, Engelke T, Meyer HE, Heber M, et al.

J Allergy Clin Immunol, 1993 Nov;92(5):690-7.
PMID: 8227860

BACKGROUND: Allergy to latex-containing articles is becoming more and more important because it can result in unexpected life-threatening anaphylactic reactions in sensitized individuals.
METHODS: A protein of 58 kd with an isoelectric point of 8.45 was purified from raw latex and from latex gloves and identified as the major allergen, completely blocking specific IgE antibodies in the serum of latex-sensitized subjects. The allergen is a noncovalent homotetramer molecule, in which the 14.6 kd monomer was identified, by amino acid composition and sequence homologies of tryptic peptides, to be the rubber elongation factor found in natural latex of the Malaysian rubber tree.
RESULTS: Competitive immunoinhibition tests showed that the starch powder covering the finished gloves is the airborne carrier of the allergen, resulting in bronchial asthma on inhalation. The purified allergen can induce allergic reactions in the nanogram range.
CONCLUSION: The identification of the allergen (Hev b I) may help to eliminate it during the production of latex-based articles in the future.

Matched MeSH terms: Electrophoresis, Polyacrylamide Gel
Electrophoretic detection of salivary alpha-amylase activity

Rahim ZH, Yaacob HB

J Nihon Univ Sch Dent, 1992 Dec;34(4):273-7.
PMID: 1283755

Fresh samples of human whole saliva containing approximately 20-40 micrograms protein were analyzed using SDS-polyacrylamide slab gel electrophoresis systems. More than 20 protein bands were revealed by Coomassie Brilliant Blue R 250 staining. Some of the protein bands were shown to be glycoprotein-positive with PAS (periodic acid-Schiff) reagent. The protein bands with alpha-Amylase activity appeared within a molecular weight range of 120,000-180,000, which is 2 to 2.8 times higher than the normal molecular weight reported for alpha-Amylase from parotid saliva, and showed positive staining with PAS reagent. These results show that the alpha-Amylase in whole saliva appears to exist in a macromolecular form which is not dissociated in the presence of sodium dodecyl sulfate (SDS).

Matched MeSH terms: Electrophoresis, Polyacrylamide Gel
Purification and characterization of two acidic phospholipase A2 enzymes from king cobra (Ophiophagus hannah) snake venom

Tan NH, Saifuddin MN

Int. J. Biochem., 1990;22(5):481-7.
PMID: 2347427

1. The two major phospholipase A2 enzymes (OHPLA-DE1 and OHPLA-DE2) of king cobra (Ophiophagus hannah) venom have been purified to electrophoretic homogeneity. 2. The isoelectric points of OHPLA-DE1 and OHPLA-DE2 were 3.81 and 3.89, respectively and the Mws were 14,000 and 15,000, respectively, as estimated by Sephadex G-75 gel filtration chromatography; and 14,000 as estimated by SDS-PAGE. 3. The enzymes were not lethal to mice at a dosage of 10 micrograms/g body wt by i.v. route. Both phospholipase A2 enzymes, however, exhibited moderate edema-inducing and anti-coagulant activities. 4. Bromophenacylation of the enzymes reduced the enzymatic activity drastically but did not affect the edema-inducing activity of the enzymes.

Matched MeSH terms: Electrophoresis, Polyacrylamide Gel
Isolation and characterization of the major phospholipase A2 from the venom of Trimeresurus purpureomaculatus (shore pit viper)

Nget Hong Tan, Chon Seng Tan, Hun Teck Khor

Int. J. Biochem., 1989;21(12):1421-6.
PMID: 2612728

1. The major phospholipase A2 (PLA-DE4) of the venom of Trimeresurus purpureomaculatus (shore pit viper) has been purified to electrophoretic homogeneity. 2. The isoelectric point of the purified enzyme was determined to be 4.20, and the mol. wt was 31,700 as estimated by Sephadex G-75 gel filtration chromatography; and 14,000 as estimated by SDS-polyacrylamide gel electrophoresis. The purified enzyme hydrolyzed phosphatidylcholine (PC) faster than phosphatidylethanolamine (PE), whereas phosphatidylserine (PS) was not hydrolyzed at all (PC greater than PE greater than PS =0). However, in reaction system consisted of mixtures of PC and PS, phosphatidylserine was effectively hydrolyzed by the enzyme. 4. The phospholipase A2 exhibited edema-forming activity but not hemolytic, hemorrhagic or anticoagulant activities. It was not lethal to mice at a dosage of 10 micrograms/g by i.v. route.

Matched MeSH terms: Electrophoresis, Polyacrylamide Gel
Iron regulated outer membrane proteins (IROMPs) as potential targets against carbapenem-resistant Acinetobacter spp. isolated from a Medical Centre in Malaysia

Hwa WE, Subramaniam G, Mansor MB, Yan OS, Gracie, Anbazhagan D, et al.

Indian J Med Res, 2010 Apr;131:578-83.
PMID: 20424311

Carbapenem-resistant Acinetobacter spp. have gained increasing significance as opportunistic pathogens in hospitalized patients. Carbapenem resistance is often associated with the loss and/or decrease in outer membrane proteins (OMP) and overexpression of multidrug efflux systems. However, carbapenem-hydrolysing beta-lactamases of Ambler Class B (metallo-enzymes) and Ambler Class D (oxacillinases) have also been detected in Acinetobacter spp. In this study we have investigated the role of the iron regulated outer membrane protein (IROMPs) and the loss of a 29-kDa OMP in carbapenem resistance of Acinetobacter calcoaceticus.

Matched MeSH terms: Electrophoresis, Polyacrylamide Gel
Fulltext Lateral flow test using Echinococcus granulosus native antigen B and comparison of IgG and IgG4 dipsticks for detection of human cystic echinococcosis

Khalilpour A, Sadjjadi SM, Moghadam ZK, Yunus MH, Zakaria ND, Osman S, et al.

Am J Trop Med Hyg, 2014 Nov;91(5):994-9.
PMID: 25200268 DOI: 10.4269/ajtmh.14-0170

Cystic echinococcosis (CE) caused by infection with Echinococcus granulosus is of major concern for humans in many parts of the world. Antigen B was prepared from E. granulosus hydatid fluid, and Western blots confirmed eight batches showing a band corresponding to the 8-/12-kDa subunit with positive serum and no low-molecular mass band (< 15 kDa) with negative serum. The batches were pooled and used to prepare lateral flow immunoglobulin G4 (IgG4) and IgG dipsticks. Diagnostic sensitivity was determined using serum samples from 21 hydatidosis patients, and diagnostic specificity was established using sera from 17 individuals infected with other parasites and 15 healthy people. IgG4 dipstick had a diagnostic sensitivity of 95% (20 of 21) and a specificity of 100% (32 of 32). The IgG dipstick had a sensitivity of 100% (21 of 21) and a specificity of 87.5% (28 of 32). Thus, both IgG and IgG4 dipsticks had high sensitivities, but IgG4 had greater specificity for the diagnosis of human CE.

Matched MeSH terms: Electrophoresis, Polyacrylamide Gel
Fulltext Feasibility of T-cell receptor gamma (TCRgamma) gene rearrangement on formalin-fixed, paraffin-embedded tissues by PCR assays

Tai YC, Peh SC

Singapore Med J, 2003 May;44(5):250-5.
PMID: 13677361

T- and B-lymphocytes are involved in recognition of foreign antigen by the specificity of their surface T-cell receptor and immunoglobulin, generated by gene rearrangement. Each T- and B-lymphocyte carries unique rearranged TCR or immunoglobulin gene, which has been applied to detect clonal from non-clonal T- and B-cell proliferation.

Matched MeSH terms: Electrophoresis, Polyacrylamide Gel
Fulltext Isolation, Purification, and Characterization of Heparinase from Streptomyces variabilis MTCC 12266

Singh V, Haque S, Kumari V, El-Enshasy HA, Mishra BN, Somvanshi P, et al.

Sci Rep, 2019 04 24;9(1):6482.
PMID: 31019210 DOI: 10.1038/s41598-019-42740-7

Arterial/venous thrombosis is the major cardiovascular disorder accountable for substantial mortality; and the current demand for antithrombotic agents is extensive. Heparinases depolymerize unfractionated heparin (UFH) for the production of low molecular-weight heparins (LMWHs; used as anticoagulants against thrombosis). A microbial strain of Streptomyces sp. showing antithrombotic activity was isolated from the soil sample collected from north India. The strain was characterized by using 16S rRNA homology technique and identified as Streptomyces variabilis MTCC 12266 capable of producing heparinase enzyme. This is the very first communication reporting Streptomyces genus as the producer of heparinase. It was observed that the production of intracellular heparinase was [63.8 U/mg protein (specific activity)] 1.58 folds higher compared to extracellular heparinase [40.28 U/mg protein]. DEAE-Sephadex A-50 column followed by Sepharose-6B column purification of the crude protein resulted 19.18 folds purified heparinase. SDS-PAGE analysis of heparinase resulted an estimated molecular-weight of 42 kDa. It was also found that intracellular heparinase has the ability to depolymerize heparin to generate LMWHs. Further studies related to the mechanistic action, structural details, and genomics involved in heparinase production from Streptomyces variabilis are warranted for large scale production/purification optimization of heparinase for antithrombotic applications.

Matched MeSH terms: Electrophoresis, Polyacrylamide Gel
Fulltext Assessing SABU (Serum Anti Bisa Ular), the sole Indonesian antivenom: A proteomic analysis and neutralization efficacy study

Tan CH, Liew JL, Tan KY, Tan NH

Sci Rep, 2016 11 21;6:37299.
PMID: 27869134 DOI: 10.1038/srep37299

Serum Anti Ular Bisa (SABU) is the only snake antivenom produced locally in Indonesia; however, its effectiveness has not been rigorously evaluated. This study aimed to assess the protein composition and neutralization efficacy of SABU. SDS polyacrylamide gel electrophoresis, size-exclusion liquid chromatography and shotgun proteomics revealed that SABU consists of F(ab')2 but a significant amount of dimers, protein aggregates and contaminant albumins. SABU moderately neutralized Calloselasma rhodostoma venom (potency of 12.7 mg venom neutralized per ml antivenom, or 121.8 mg venom per g antivenom protein) and Bungarus fasciatus venom (0.9 mg/ml; 8.5 mg/g) but it was weak against the venoms of Naja sputatrix (0.3 mg/ml; 2.9 mg/g), Naja sumatrana (0.2 mg/ml; 1.8 mg/g) and Bungarus candidus (0.1 mg/ml; 1.0 mg/g). In comparison, NPAV, the Thai Neuro Polyvalent Antivenom, outperformed SABU with greater potencies against the venoms of N. sputatrix (0.6 mg/ml; 8.3 mg/g), N. sumatrana (0.5 mg/ml; 7.1 mg/g) and B. candidus (1.7 mg/ml; 23.2 mg/g). The inferior efficacy of SABU implies that a large antivenom dose is required clinically for effective treatment. Besides, the antivenom contains numerous impurities e.g., albumins that greatly increase the risk of hypersensitivity. Together, the findings indicate that the production of SABU warrants further improvement.

Matched MeSH terms: Electrophoresis, Polyacrylamide Gel
Fulltext Characterization and purification of protease extracted from two maturity stages of ‘noni’ (Morinda citrifolia L.) fruit

Normah Ismail, Nurulain Abd Razak

Scientific Research Journal, 2014;11(2):0-0.
MyJurnal

Protease was extracted from two maturity stages of noni fruits (Morinda citrifolia L.), unripe (stage 1) and ripe (stage 5). The crude extract was partially purified by acetone precipitation method followed by dialysis, gel filtration chromatography and freeze drying. Protein concentrations, proteolytic activity, molecular weight distribution, pH stability, temperature stability and storage efficiency of the resulting protease were evaluated. The unripe and ripe noni fruit contains 0.65 and 0.35% protein, respectively. Molecular weight of the proteases from both stages ranged approximately between 3 to 28 kDa based on the SDS-PAGE results. The optimum activity were at pH 7s and 6, temperatures of 40 and 50°C, respectively for proteases obtained from the unripe and ripe fruit. Analysis from the freeze dried protease indicated that protease from ripe noni fruits had higher protein concentration and specific activity compared to those from unripe fruit. However, it is more sensitive to pH and temperature and less stable during storage as it shows lower proteolytic activity compared to protease from unripe fruit. Based on its high proteolytic activity reaching up to 70.31 U/mg and storage stability (30% lost of activity), noni fruit could be an alternative source of plant protease.

Matched MeSH terms: Electrophoresis, Polyacrylamide Gel
Fulltext Effects of precipitation methods on the properties of protease extracted from starfruit (Averrhoa carambola L.) of different maturity index

Normah, Ismail, Ezzana Zuraini, Zainuddin

Scientific Research Journal, 2015;12(1):1-12.
MyJurnal

Proteases were extracted from starfruit at maturity Index 2 (unripe, light green) and Index 7 (very ripe, orange) and partially purified using acetone and 40% ammonium sulfate precipitations. Higher yield and proteolytic activity were observed for proteases purified using acetone than 40% ammonium sulfate. As for maturity index, yield and protein concentration of proteases from Index 2 were higher than those from Index 7. SDS-PAGE result showed intense bands for acetone proteases while a distinct band at 50 kDa was observed in all the proteases. Enzyme activity decreased during the seven days storage at 4°C with minimum relative activity of 70% achieved for acetone proteases at day seven. This study suggested that acetone precipitation is more effective method for purifying starfruit protease based on the yield and proteolytic activity compared to using 40% ammonium sulphate precipitation. In order to obtain higher protein concentration and proteolytic activity, starfruit at the unripe stage, Index 2 is a better raw material than Index 7 to be used for protease production.

Matched MeSH terms: Electrophoresis, Polyacrylamide Gel
Proteomic analysis of stored core oil palm trunk (COPT) sap identifying proteins Related to stress, disease resistance and differential gene/protein expression

Marhaini Mostapha, Noorhasmiera Abu Jahar, Sarani Zakaria, Sharifah Nabihah Syed Jaafar, Kamalrul Azlan Azizan, Wan Mohd Aizat

Sains Malaysiana, 2018;47:1259-1268.

Oil palm is the major crop grown and cultivated in various Asian countries such as Malaysia, Indonesia and Thailand.
The core of oil palm trunk (COPT) consists of high sugar content, hence suitable for synthesis of fine chemicals and
biofuels. Increase of sugar content was reported previously during prolonged COPT storage. However, until now, there
has been no report on protein profiles during storage. Therefore, in this study, protein expression of the COPT during the
storage period of one to six weeks was investigated using sodium dodecyl sulphate polyacrylamide gel electrophoresis
(SDS-PAGE) coupled with optical density quantification and multivariate analyses for measuring differentially expressed
proteins. Accordingly, protein bands were subjected to tryptic digestion followed by tandem mass spectrometry (nanoLCMS/MS)
protein identification. The results from SDS-PAGE showed consistent protein bands appearing across the biological
replicates ranging from 10.455 to 202.92 kDa molecular weight (MW) regions. The findings from the principal component
analysis (PCA) plot illustrated the separation pattern of the proteins at weeks 4 and 5 of storage, which was influenced
mainly by the molecular weights of 14.283, 25.543, 29.757, 30.549, 31.511, 34.585 and 84.395 kDa, respectively. The
majority of these proteins are identified as those involved in stress- and defense-related, disease resistance, as well
as gene/protein expression processes. Indeed, these proteins were mostly upregulated during the later storage period
suggesting that long-term storage may influence the molecular regulation of COPT sap.

Matched MeSH terms: Electrophoresis, Polyacrylamide Gel
Optimization of Enzymatic Hydrolysis of tilapia muscle (Oreochromis niloticus) using Response Surface Methodology (RSM)

Jumardi Roslan, Siti Mazlina Mustapa Kamal, Khairul Faezah Md. Yunos, Norhafizah Abdullah

Sains Malaysiana, 2014;43:1715-1723.

Fish protein hydrolysate was prepared from tilapia muscle using commercial Alcalase enzyme. Optimization of enzymatic hydrolysis process for preparing tilapia muscle protein hydrolysates (TMPH) was performed by employing central composite design (CCD) method of response surface methodology (RSM). O-phtaldialdehyde (OPA) method was employed to calculate the degree of hydrolysis (DH), which is the key parameter for monitoring the reaction of protein hydrolysis. The suggested model equation was proposed based on the effects of pH, temperature, substrate concentration and enzyme concentration on the DH. Optimum enzymatic hydrolysis conditions using Alcalase enzyme were obtained at pH7.5, temperature of 50oC, substrate concentration of 2.5% and enzyme concentration of 4.0%. Under these conditions, the highest value of the DH was achieved at 25.16% after hydrolysing at 120 min. The TMPH was further assessed for their nutritional value with respect to chemical and amino acid compositions. Molecular weight distributions of TMPH were characterized by SDS-PAGE. TMPH contains moderate amount of protein (28.14%) and good nutritive value with respect to the higher total amino acid composition (267.57 mg/g). Glutamic acid, aspartic acid and lysine were the most abundant amino acids present in TMPH with values 42.68, 29.16 and 26.21 mg/g, respectively. Protein hydrolysates from tilapia muscle containing a desirable peptide with low molecular weight which may potentially to be used as functional food products.

Matched MeSH terms: Electrophoresis, Polyacrylamide Gel
Infeksi Plasmodium berghei dan kesannya ke atas pengisyaratan MAP kinase eritrosit perumah

Mohd Fakharul Zaman Raja Yahya, Hasidah Mohd Sidek

Sains Malaysiana, 2009;38.

Kajian ini melibatkan pemantauan perkembangan parasitemia dan taburan morfologi Plasmodium berghei sewaktu infeksi parasit dalam mencit, serta penentuan kesan infeksi P. berghei ke atas pengisyaratan MAP kinase eritrosit perumah. Analisis mikroskop ke atas slaid calitan darah terwarna-Giemsa yang disediakan daripada mencit terinfeksi-P. berghei (strain PZZ1/00) menunjukkan darjah parasitemia mencapai sehingga 70% dalam masa dua minggu selepas penyuntikan parasit. Morfologi cecincin dan trofozoit parasit dicerap dengan jelas sepanjang tempoh infeksi manakala morfologi skizon parasit hanya dicerap dengan ketara selepas hari ketiga selepas penyuntikan parasit. Pemblotan Western [antibodi primer: anti-MAP kinase (ERK-1/2 tak terfosfat) monoklon; antibodi sekunder: anti-IgG, poliklon terkonjugat-HRP] ke atas protein sitosol eritrosit terinfeksi-P. berghei (70% parasitemia) susulan pemisahan SDS-PAGE menunjukkan bahawa keamatan protein imunoreaktif-MAP kinase eritrosit berberat molekul 42 dan 44 kDa didapati meningkat secara signifikan (p<0.05) pada 70% iaitu peningkatan sebanyak 21.5% dan 22.3% masing-masing berbanding sampel kawalan tanpa infeksi. Samada kesan infeksi P. berghei (70% parasitemia) ke atas pengisyaratan MAP kinase perumah ini berkaitan dengan pengaktifan enzim ini perlu dikaji dengan lebih lanjut.

Matched MeSH terms: Electrophoresis, Polyacrylamide Gel
Evaluation of bovine antibody responses to haemorrhagic septicaemia vaccine

Johnson RB, Dawkins HJ, Spencer TL, Saharee AA, Bahaman AR, Ramdani, et al.

Res Vet Sci, 1989 Sep;47(2):277-9.
PMID: 2508206

ELISA and immunoblotting techniques were used to examine the humoral immune response to Pasteurella multocida, in bovine sera from Indonesia and Malaysia. Elevated levels of antibody to a crude lipopolysaccharide preparation were found in vaccinated animals. In addition to the response to lipopolysaccharide, antibodies from the vaccinated cattle strongly labelled five to six of the 40 protein bands in this organism.

Matched MeSH terms: Electrophoresis, Polyacrylamide Gel
Secretory expression in Escherichia coli and single-step purification of a heat-stable alkaline protease

Fu Z, Hamid SB, Razak CN, Basri M, Salleh AB, Rahman RN

Protein Expr Purif, 2003 Mar;28(1):63-8.
PMID: 12651108

Bacteriocin release proteins (BRPs) can be used for the release of heterologous proteins from the Escherichia coli cytoplasm into the culture medium. The gene for a highly thermostable alkaline protease was cloned from Bacillus stearothermophilus F1 by the polymerase chain reaction. The recombinant F1 protease was efficiently excreted into the culture medium using E. coli XL1-Blue harboring two vectors: pTrcHis bearing the protease gene and pJL3 containing the BRPs. Both vectors contain the E. coli lac promoter-operator system. In the presence of 40 microM IPTG, the recombinant F1 protease and the BRP were expressed and mature F1 protease was released into the culture medium. This opens the way for the large-scale production of this protease in E. coli. The recombinant enzyme was purified through a one-step heat treatment at 70 degrees C for 3h and this method purified the protease to near homogeneity. The purified enzyme showed a pH optimum of 9.0, temperature optimum of 80 degrees C, and was stable at 70 degrees C for 24h in the pH range from 8.0 to 10.0. The enzyme exhibited a high degree of thermostability with a half-life of 4 h at 85 degrees C, 25 min at 90 degrees C, and was inhibited by the serine protease inhibitor phenylmethylsulfonyl fluoride (PMSF).

Matched MeSH terms: Electrophoresis, Polyacrylamide Gel

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