Production and Characterization of Monoclonal Antibody against Vit v1: A Grape Allergen Belonging to Lipid Transfer Protein Family

Allergy to non-specific lipidtransfer protein (nsLTP), the major allergen of grape (Vit v1), is considered as one of the most common fruit allergies in Iran. Therefore, a specific monoclonal antibody (mAb) can be used for the characterization and assessment of. Accordingly, this study aimed to generate and characterize a mAb against Vit v1 with a diagnostic purpose. To this end, Vit v1 allergen (9 kDa) was extracted using a modified Bjorksten extraction method. Natural Vit v1-immunized mouse splenocytes were fused with SP2/0Ag-14 myeloma cells for generating hybridoma cells. Specific antibody-secreting Hybridoma cells were selected using ELISA. Finally, anti-Vit v1 mAb was characterized by western blotting, ELISA, and isotyping methods. In the current study, a 9 kDa (Vit v1) protein was attained fromcrude and fresh juice of grape extracts and the isotype of desired anti-Vit v1 mAb was determined as IgM with K light chain. In addition, The ELISA results demonstrated that anti-Vit v1 mAb was specified against natural Vit v1 in the grape cultivar and related LTP allergens, such as Pla or 3 (p<0.0001). In the present study, a specific mAb was produced for detecting the LTP allergen. This mAb with a confirmed specificity can be utilized for evaluating the LTP allergens and their allergenicity in different grape cultivars. © 2020 Tehran University of Medical Science

Measurement of melatonin in body fluids: Standards, protocols and procedures

Abstract: The circadian rhythm of melatonin in saliva or plasma, or of the melatonin metabolite 6‐ sulphatoxymelatonin in urine, is a defining feature of suprachiasmatic nucleus function, the endogenous oscillatory pacemaker. These measurements are useful to evaluate problems related to the onset or offset of sleep and for assessing phase delays or advances of rhythms in entrained individuals. Additionally, they have become an important tool for psychiatric diagnosis, its use being recommended for phase typing in patients suffering from sleep and mood disorders. Thus, the development of sensitive and selective methods for the precise detection of melatonin in tissues and fluids of animals emerges as necessary. Due to its low concentration and the co‐existence of many other endogenous compounds in blood, the determination of melatonin has been an analytical challenge. This review discusses current methodologies employed for detection and quantification of melatonin in biological fluids and tissues

Cardioprotective effects of Achillea wilhelmsii on the isolated rat heart in ischemia–reperfusion

Context: There are some reports about protective effects of Achillea on the heart. Objective: We investigated the effect of Achillea wilhelmsii extract on cardiac function during ischemia–reperfusion (I/R) injury in the isolated rat heart. Materials and methods: 60 male Wistar rats were randomly divided into 6 groups; 1: Control group, 2: Control-ischemia (CI) 3: vitamin C (10 mg/kg), 4–6: Extract groups (E 100, E 200 and E 400 mg/kg). The animals received normal saline, vitamin C or A. wilhelmsii extract orally for 4 weeks. At the end of the treatment, the hearts were subjected to in vitro I/R Injury (20 min of global ischemia, followed by 40 min of reperfusion, Langendorff's mode). Heart rate (HR) and left ventricular pressure (LVP) were measured using a pressure transducer connected to a data acquisition system. Lactate dehydrogenase (LDH) and creatine kinase (CK) activities in the effluent were measured to determine the myocardial injury degree. The malondialdehyde (MDA), total thiol groups (-SH), superoxide anion dismutase (SOD) and catalase (CAT) in myocardial tissue were detected to determine the oxidative stress degree. Results: Pretreatment with Achillea wilhlemsii significantly decreased the LDH, CK activities, and MDA level, while it increased the LVDP, ±dp/dtmax, rate-pressure product (RPP), SH groups, SOD and CAT activities, and also the coronary artery flow. Discussion and conclusion: Our findings indicated that Achillea wilhelmsii could provide protection for heart against the I/R injury which may be related to the improvement of myocardial oxidative stress states

Contribution of estradiol in sex-dependent differences of pentylenetetrazole-induced seizures in rats

In the present study the contribution of estradiol in sex-dependent differences of pentylenetetrazole (PTZ)-induced seizures was investigated in rats. The rats were divided into four groups: 1) sham, 2) ovariectomized (OVX), 3) ovariectomized-estradiol (OVX-Est) and 4) male. The OVX-Est group received estradiol valerate (2 mg/kg; i.m/4 weeks) while, male, sham and OVX groups received vehicle. The animals were injected by PTZ (90 mg/kg). The latencies to minimal clonic seizures (MCS) and generalized tonic-clonic seizures (GTCS), were recorded. Serum 17β-estradiol and testosterone levels were also determined using an Elisa kit. GTCS latency in OVX rats was higher than in sham-operated animals (P < 0.05). MCS and GTCS latency in the male group was significantly higher than in the sham, OVX and OVX-Est groups (P < 0.001 and P < 0.01). There was no significant difference in MCS or GTCS latencies among OVX-Est, sham and OVX groups. Serum 17β-estradiol level in the OVX group was significantly lower than in the sham (P < 0.01) and in the OVX-Est group it was higher than in the sham, OVX and male groups (P < 0.01 and P < 0.001). Serum testosterone level in the male group was significantly higher than in all the other three groups (P < 0.001). It seems that testosterone probably has a more efficient role than estradiol in the gender dependent difference in seizure caused by PTZ in rats

A Novel Monoclonal Antibody Against A60 Antigen of Mycobacterium bovis Bacillus Calmette-Guerin

Mycobacterium contains several immunologically active substances, which play a principal role in mycobacterial diseases. The majority of the highly antigenic proteins present in mycobacterial homogenates are components of the A60 complex. In this study, A60 antigen was prepared from cytoplasm of Mycobacterium bovis Bacillus Calmette-Guerin (BCG). Cytoplasm was fractionated by passage through the column of sepharose 6B and ConA-sepharose 4B. After purification of spleen cells of the immunized mice, the cells were fused with SP2/0 myeloma cells. Four clone cell lines producing antibody against A60 antigens were established and each clone was tested for immunoreactivity against purified A60 by ELISA and immunoblotting. The clone designated DEB7 reacted strongly with A60. Immunoblotting using MAb DEB7 showed that this MAb binds to a single protein of A60 subunit with a molecular weight of 65 kDa. This subunit of A60 M. bovis recognized by DEB7 MAb could be used to increase the sensitivity and specificity of immunoassay or other potential roles in mycobacterium infection

Evaluation of micronuclei and antioxidant status in hospital radiation workers occupationally exposed to low-dose ionizing radiation

Abstract Purpose There is scientific evidence that ionizing radiation (IR) can be responsible for various health hazards that are one of the concerns in occupational exposure. This study was performed to evaluate DNA damage and antioxidant status in hospital workers who are occupationally exposed to low doses of IR. Materials and methods In this study, twenty occupationally exposed to low doses of IR (CT and angiography) comprising with control groups which matched them. In order to investigate the effects of chronic irradiation of radiation workers, Micronuclei (MN) frequency and the antioxidant activity of Superoxide Dismutase (SOD), Catalase (CAT) and Total Antioxidant Capacity (TAC) were measured. Then, to check adaptation against high challenge dose, the samples (in all groups) were irradiated in vitro and MN frequency was compared. Finally, to investigated the effect of the high dose after the acute and chronic low dose of ionizing radiation, MN frequency was compared in two groups (the control group that was to in-vitro irradiated (acute low dose + high dose) and radiation workers (chronic low dose + high dose)). Results MN frequency in the occupationally exposed group (n = 30) increased significantly when compared to the control group (p-value  0.05). Conclusions We observed that exposure to low doses of IR leads to increased cytogenetic damage, could not cause an adaptive-response, and improve antioxidant capacity in radiation workers. Controlling healthcare workers' exposure is the first step to improving the health of hospital workers and the quality of patient care, thus decreasing human and economic costs