Optimization of Expression and Purification of Recombinant Mouse plac1

Background: Placenta-specific 1 (PLAC1) is one of the recently-discovered Cancer-Testis-Placenta (CTP) antigen with restricted normal tissue and ectopic expression in a wide range of cancer cells from different histological origins. The production of recombinant human PLAC1 has already been optimized; however, no study has been reported so far on the production and purification of mouse plac1. In this study, mouse plac1 expression and purification was optimized in a prokaryotic system and the effects of the generated proteins on inducing humoral responses in mice were investigated. Methods: A fusion protein containing full extracellular domain of mouse plac1, immunostimulatory peptides, tetanus toxin P2P30 and PADRE and KDEL3 signal (main plac1), and the same fragment without immunostimulatory peptides (control plac1) was produced. To optimize production and purification steps, different parameters including bacterial strain, cultivation temperature, cultivation time, IPTG concentration, culture medium, and also different buffers for purification of the recombinant proteins were tested. After confirming the identity of recombinant plac1 proteins with Western Blotting (WB) and ELISA assays, these proteins were subcutaneously injected in mice with Freund's adjuvant and the anti-plac1 antibody response was detected by ELISA. Results: The optimal expression level of main and control plac1 was obtained in BL21 (DE3) and TB culture medium in the presence of 0.25 mM IPTG after 24 hr of induction at 15°C. The buffer containing 2% sarkosyl produced higher yield and purity. Our results showed specific reactivity of anti-human recombinant plac1 polyclonal antibody with both main and control plac1 recombinant proteins in WB and ELISA analysis. Both proteins induced humoral responses in mice; however, anti-plac1 antibody titer was significantly higher in sera of mice immunized with main compared to control plac1. Conclusion: In this study, an optimized protocol for production and purification of mouse plac1 was reported and it was shown that insertion of immunostimulatory peptides in gene construct could efficiently enhance humoral immune responses against mouse plac1, which could potentially augment cellular immune responses against plac1 leading to more effective anti-cancer responses.

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Effect of Resveratrol Dry Suspension on Immune Function of Piglets

Evidence-based Complementary and Alternative Medicine ◽

10.1155/2018/5952707 ◽

2018 ◽

Vol 2018 ◽

pp. 1-10 ◽

Cited By ~ 6

Author(s):

Qiuting Fu ◽

Qiankun Cui ◽

Yi Yang ◽

Xinghong Zhao ◽

Xu Song ◽

...

Keyword(s):

Immune Responses ◽

Immune Function ◽

Foot And Mouth Disease ◽

Classical Swine Fever ◽

Sod Activity ◽

Humoral And Cellular Immunity ◽

Humoral Immune ◽

Humoral Immune Responses ◽

Mouth Disease Virus ◽

Wide Range

Resveratrol, a polyphenolic plant antitoxin, has a wide range of pharmacological activities. In this study, we systematically evaluated the effects of resveratrol dry suspension (RDS) on immune function in piglets that were treated with different doses of RDS for 2 weeks. The results showed that the RDS has significant effects on the development, maturation, proliferation, and transformation of T lymphocytes. RDS could regulate humoral immune responses by upregulating the release of IFN-γ and downregulating the release of TNF-α. After piglets were vaccinated against classical swine fever virus and foot-and-mouth disease virus, the antibody titers were significantly increased. RDS treatment showed an excellent resistance to enhance T-SOD activity. Values of blood routine and blood biochemistry showed no toxicity. These results suggested that RDS could be considered as an adjuvant to enhance immune responses to vaccines, as well as dietary additives for animals to enhance humoral and cellular immunity.

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Overcoming Immunity to a Viral Vaccine by DNA Priming before Vector Boosting

Journal of Virology ◽

10.1128/jvi.77.1.799-803.2003 ◽

2003 ◽

Vol 77 (1) ◽

pp. 799-803 ◽

Cited By ~ 161

Author(s):

Zhi-yong Yang ◽

Linda S. Wyatt ◽

Wing-pui Kong ◽

Zoe Moodie ◽

Bernard Moss ◽

...

Keyword(s):

Immune Responses ◽

Viral Vectors ◽

Ebola Virus ◽

Viral Vector ◽

Hemorrhagic Fever ◽

Viral Immunity ◽

Humoral Immune ◽

Humoral Immune Responses ◽

Viral Vaccine ◽

Humoral Responses

ABSTRACT Replication-defective adenovirus (ADV) and poxvirus vectors have shown potential as vaccines for pathogens such as Ebola or human immunodeficiency virus in nonhuman primates, but prior immunity to the viral vector in humans may limit their clinical efficacy. To overcome this limitation, the effect of prior viral exposure on immune responses to Ebola virus glycoprotein (GP), shown previously to protect against lethal hemorrhagic fever in animals, was studied. Prior exposure to ADV substantially reduced the cellular and humoral immune responses to GP expressed by ADV, while exposure to vaccinia inhibited vaccine-induced cellular but not humoral responses to GP expressed by vaccinia. This inhibition was largely overcome by priming with a DNA expression vector before boosting with the viral vector. Though heterologous viral vectors for priming and boosting can also overcome this effect, the paucity of such clinical viral vectors may limit their use. In summary, it is possible to counteract prior viral immunity by priming with a nonviral, DNA vaccine.

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Limited ability of humoral immune responses in control of viremia during infection with SIVsmmD215 strain

Blood ◽

10.1182/blood-2008-09-177741 ◽

2009 ◽

Vol 113 (18) ◽

pp. 4250-4261 ◽

Cited By ~ 25

Author(s):

Thaidra Gaufin ◽

Rajeev Gautam ◽

Melissa Kasheta ◽

Ruy Ribeiro ◽

Erin Ribka ◽

...

Keyword(s):

Immune Responses ◽

Antibody Production ◽

Neutralizing Antibody ◽

Viral Loads ◽

Humoral Immune ◽

Humoral Immune Responses ◽

Significant Difference ◽

Anti Cd20 ◽

And Control ◽

The Impact

AbstractWe investigated the impact of rhesus macaque (RM) B-cell depletion before inoculation with the isolate SIVsmmD215. Seven RMs were treated every 3 weeks with 50 mg/kg of an anti-CD20 antibody (rituximab) starting 7 days before inoculation for 2 (n = 4) and 5 (n = 3) months. Four control animals received no antibody. Three animals were completely depleted of CD20+ B cells, but 4 were only partially depleted of CD20 cells in the LNs and intestine. The decrease in antibody production was consistent with the efficacy of tissue CD20 depletion. Seroconversion and neutralizing antibody production was significantly delayed in animals showing complete tissue CD20 depletion and remained at low titers in all CD20-depleted RMs. Surprisingly, there was no significant difference in acute or chronic viral loads between CD20-depleted and control animal groups. There was a tendency for lower viral set points in CD20-depleted animals. At 6 weeks after inoculation, cellular immune responses were significantly stronger in CD20-depleted animals than in controls. There was no significant difference in survival between CD20-depleted and control animals. Our data suggest that a deficiency of Ab responses did not markedly affect viral replication or disease progression and that they may be compensated by more robust cellular responses.

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Humoral Immune Responses against the Mycobacteriumtuberculosis 38-Kilodalton, MTB48, and CFP-10/ESAT-6 Antigens in Tuberculosis

Clinical and Vaccine Immunology ◽

10.1128/cvi.00287-09 ◽

2010 ◽

Vol 17 (3) ◽

pp. 372-375 ◽

Cited By ~ 35

Author(s):

Xueqiong Wu ◽

Yourong Yang ◽

Junxian Zhang ◽

Bangying Li ◽

Yan Liang ◽

...

Keyword(s):

Immune Responses ◽

Healthy Subjects ◽

Enzyme Linked Immunosorbent Assay ◽

Humoral Immune ◽

Humoral Immune Responses ◽

Single Antigen ◽

Human Sera ◽

Low Sensitivity ◽

Humoral Responses ◽

Culture Negative

ABSTRACT The diagnosis of smear-negative and culture-negative patients with active tuberculosis (TB) is challenging. The detection of Mycobacterium tuberculosis-specific antibodies in human sera has been an important diagnostic aid. However, detection of antibody responses to a single antigen usually has a low sensitivity for diagnosis of TB. In this study, humoral immune responses against recombinant M. tuberculosis 38-kDa, MTB48, and CFP-10/ESAT-6 (culture filtrate protein 10/6-kDa early secreted antigen target of M. tuberculosis) antigens in 250 Chinese TB patients and 260 healthy subjects were evaluated by an enzyme-linked immunosorbent assay (ELISA). The levels of antibodies against those antigens in TB patients, even in bacterium-negative ones, were significantly higher than those in healthy subjects (P < 0.001). The serodiagnostic sensitivities to detect antibodies against individual antigens, i.e., recombinant M. tuberculosis 38-kDa, MTB48, and CFP-10/ESAT-6 antigens, in TB patients were 73.6%, 73.2%, and 60.4%, respectively, with specificities of 85.4%, 77.7%, and 73.8%, respectively. Importantly, the sensitivity to positively detect humoral responses to one of the antigens increased further. Our data suggest that the humoral immune responses to M. tuberculosis antigens in TB patients are heterogeneous. The 38-kDa, MTB48, and CFP-10/ESAT-6 antigens can be used as the cocktail antigens in the serodiagnosis of active TB, especially for smear- or culture-negative TB cases.

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Longitudinal immune dynamics of mild COVID-19 define signatures of recovery and persistence

10.1101/2021.05.26.442666 ◽

2021 ◽

Author(s):

Aarthi Talla ◽

Suhas V Vasaikar ◽

Maria P Lemos ◽

Zoe Moodie ◽

Mark-Phillip Lee Pebworth ◽

...

Keyword(s):

Immune Responses ◽

Acute Infection ◽

Early Time ◽

Chromatin Accessibility ◽

Outpatient Setting ◽

Correlated Data ◽

Humoral Immune ◽

Humoral Immune Responses ◽

Serum Proteomics ◽

Humoral Responses

SARS-CoV-2 has infected over 160 million and caused more than 3 million deaths to date. Most individuals (>80%) have mild symptoms and recover in the outpatient setting, but detailed studies of immune responses have focused primarily on moderate to severe COVID-19. We deeply profiled the longitudinal immune response in individuals with mild COVID beginning with early time points post-infection (1-15 days) and proceeding through convalescence to >100 days after symptom onset. We correlated data from single cell analyses of peripheral blood cells, serum proteomics, virus-specific cellular and humoral immune responses, and clinical metadata. Acute infection was characterized by vigorous coordinated innate and adaptive activation, including an early cellular and proteomic signature that correlated with the amplitude of virus-specific humoral responses after day 30. We characterized signals associated with recovery and convalescence to define a new signature of inflammatory cytokines, gene expression, and chromatin accessibility that persists in individuals with post-acute sequelae of SARS-CoV-2 infection (PASC).

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Strong humoral immune responses against SARS-CoV-2 Spike after BNT162b2 mRNA vaccination with a sixteen-week interval between doses

10.1101/2021.09.17.21263532 ◽

2021 ◽

Author(s):

Alexandra Tauzin ◽

Shang Yu Gong ◽

Guillaume Beaudoin-Bussieres ◽

Dani Vezina ◽

Romain Gasser ◽

...

Keyword(s):

Public Health ◽

Immune Responses ◽

Vaccine Efficacy ◽

Standard Regimen ◽

Effector Functions ◽

Humoral Immune ◽

Humoral Immune Responses ◽

Health Authorities ◽

Public Health Authorities ◽

Humoral Responses

While the standard regimen of the BNT162b2 mRNA vaccine includes two doses administered three weeks apart, some public health authorities decided to space them, raising concerns about vaccine efficacy. Here, we analyzed longitudinal humoral responses including antibody binding, Fc-mediated effector functions and neutralizing activity against the D614G strain but also variants of concern and SARS-CoV-1 in a cohort of SARS-CoV-2 naive and previously infected individuals, with an interval of sixteen weeks between the two doses. While the administration of a second dose to previously infected individuals did not significantly improve humoral responses, we observed a significant increase of humoral responses in naive individuals after the 16-weeks delayed second shot, achieving similar levels as in previously infected individuals. Our results highlight strong vaccine-elicited humoral responses with an extended interval BNT162b2 vaccination for naive individuals.

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Different formulations of inactivated SARS-CoV-2 vaccine candidates in human compatible adjuvants: Potency studies in mice showed different platforms of immune responses

10.21203/rs.3.rs-1067486/v1 ◽

2021 ◽

Author(s):

Melika Haghighi ◽

Akbar khorasani ◽

Pegah Karimi ◽

Rouhollah Keshavarz ◽

Mehdi Mahdavi

Keyword(s):

Immune Responses ◽

Control Group ◽

Control Groups ◽

Humoral Immune ◽

Humoral Immune Responses ◽

Igg2a Level ◽

Specific Igg ◽

Ifn Γ ◽

And Control ◽

Ctl Activity

Abstract Several inactivated SARS-CoV-2 vaccines were approved for human use but are not highly potent. Here, different formulations of inactivated SARS-CoV-2 virus in Alum, Montanide 51VG and Montanide ISA720VG adjuvants were developed and then immune responses were assessed. SARS-CoV-2 virus was inactivated with formalin and formulated in the adjuvants. BALB/c mice were immunized subcutaneously with 4µg of experimental vaccines on days 0 and 14 and two weeks after the final immunization, IL-4 and IFN-γ cytokines, CTL activity and specific IgG titer and IgG1, IgG2a, IgG2a/IgG1 ratio and also anti-RBD IgG response were assessed. Immunization with SARS-CoV-2-Montanide ISA51VG showed a significant increase in IFN-γ cytokine versus SARS-CoV-2-Alum, SARS-CoV-2-Montanide ISA720VG and control groups (P<0.0033). Cytokine IL-4 response in SARS-CoV-2-Alum group showed a significant increase versus SARS-CoV-2-Montanide ISA51VG, SARS-CoV-2-Montanide ISA720VG and control groups (P<0.0206). In addition, SARS-CoV-2-Montanide ISA51VG vaccine induced the highest IFN-γ/IL-4 cytokine ratio versus other groups (P<0.0004). CTL activity in SARS-CoV-2- Montanide ISA51 VG and SARS-CoV-2-Montanide ISA720 VG groups showed a significant increase versus SARS-CoV-2-Alum and control groups (P<0.0075). Specific IgG titer in SARS-CoV-2- Montanide ISA51 VG and SARS-CoV-2-Montanide ISA720VG showed significant increase versus SARS-CoV-2-Alum and control groups (P<0.0143). Results of specific IgG1and IgG2a level in SARS-COV-2-Alum, SARS-COV-2-Montanide ISA51VG and SARS-COV-2-Montanide ISA720VG vaccine showed a significant increase versus the control group (P<0.0001) but SARS-COV-2-Montanide ISA51VG and SARS-COV-2-Montanide ISA 720VG groups showed highest IgG2a/IgG1 ratio and a significant increase versus SARS-COV-2-Alum group (P<0.0379). Results of anti-RBD IgG response showed that inactivated SARS-COV-2+Alum and SARS-COV-2-Montanide ISA 720VG vaccine groups demonstrated a significant increase versus SARS-COV-2-Montanide ISA51VG group. It seems that the type of vaccine formulation is a critical parameter that effect on immunologic patterns and vaccine potency. Here, results showed that human compatible oil-based adjuvants were more potent than Alum adjuvant in the induction of cellular and humoral immune responses versus SARS-CoV-2 virus.

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Enhanced Humoral Immune Responses against Toxin A and B of Clostridium difficile is Associated with a Milder Disease Manifestation

Journal of Clinical Medicine ◽

10.3390/jcm9103241 ◽

2020 ◽

Vol 9 (10) ◽

pp. 3241

Author(s):

Wasef Na’amnih ◽

Yehuda Carmeli ◽

Valeria Asato ◽

Sophy Goren ◽

Amos Adler ◽

...

Keyword(s):

Clostridium Difficile ◽

Immune Responses ◽

Large Scale ◽

Immunoglobulin A ◽

Toxin A ◽

Disease Manifestation ◽

Humoral Immune ◽

Humoral Immune Responses ◽

And Control

The role of the humoral immune response to Clostridium difficile in modulating the severity of C. difficile infection (CDI) is unclear. We compared the levels of serum immunoglobulin G (IgG) and immunoglobulin A (IgA) against toxin A (TcdA) and toxin B (TcdB) of C. difficile between CDI and control patients and according to disease severity. The levels of IgG and IgA antibodies against TcdA and TcdB were measured in sera from patients with CDI (n = 50; 19 had severe CDI) and control patients (n = 52), using ELISA. Patients with CDI had higher levels of IgG antibodies against TcdA and TcdB than controls (p = 0.001 and p = 0.04, respectively). Higher IgG levels against TcdA and TcdB were found in patients with mild vs. severe CDI 7–14 days after the diagnosis (p = 0.004 and 0.036, respectively). A factor analysis included both IgA and IgG levels against both toxins into one composite variable, which was of higher values in patients with mild vs. severe CDI (p = 0.026). In conclusion, the systemic humoral immune responses against TcdA and TcdB might modulate the severity of CDI. These preliminary findings provide a basis for future large-scale studies and support the development and evaluation of active and passive immunotherapies for CDI management.

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Humoral Immune Responses of Tuberculosis Patients in Brazil Indicate Recognition of Mycobacterium tuberculosis MPT-51 and GlcB

Clinical and Vaccine Immunology ◽

10.1128/cvi.00359-07 ◽

2008 ◽

Vol 15 (3) ◽

pp. 579-581 ◽

Cited By ~ 11

Author(s):

Cristina Melo Cardoso Almeida ◽

Arioldo C. Vasconcelos ◽

André Kipnis ◽

Ana Lúcia Andrade ◽

Ana Paula Junqueira-Kipnis

Keyword(s):

Mycobacterium Tuberculosis ◽

Immune Responses ◽

Immunosorbent Assay ◽

Immunoglobulin M ◽

Enzyme Linked Immunosorbent Assay ◽

Tuberculosis Patients ◽

Humoral Immune ◽

Humoral Immune Responses ◽

Humoral Responses

ABSTRACT The humoral responses to recombinant MPT-51 and GlcB was determined by using an enzyme-linked immunosorbent assay. Levels of immunoglobulin M (IgM) against MPT-51 and IgG against GlcB were higher among tuberculosis (TB) patients than among control individuals. When the MPT-51 and GlcB assays were combined, 90.8% specificity and 75.5% sensitivity were observed. MPT-51 and GlcB were recognized in the humoral responses of Brazilian TB patients.

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Humoral Responses and Serological Assays in SARS-CoV-2 Infections

Frontiers in Immunology ◽

10.3389/fimmu.2020.610688 ◽

2020 ◽

Vol 11 ◽

Author(s):

Yannick Galipeau ◽

Matthew Greig ◽

George Liu ◽

Matt Driedger ◽

Marc-André Langlois

Keyword(s):

Immune Responses ◽

Vaccine Development ◽

Cross Reactivity ◽

The Novel ◽

Epidemiological Models ◽

Humoral Immune ◽

Health Measures ◽

Humoral Immune Responses ◽

Humoral Responses ◽

Vaccination Campaigns

In December 2019, the novel betacoronavirus Severe Acute Respiratory Disease Coronavirus 2 (SARS-CoV-2) was first detected in Wuhan, China. SARS-CoV-2 has since become a pandemic virus resulting in hundreds of thousands of deaths and deep socioeconomic implications worldwide. In recent months, efforts have been directed towards detecting, tracking, and better understanding human humoral responses to SARS-CoV-2 infection. It has become critical to develop robust and reliable serological assays to characterize the abundance, neutralization efficiency, and duration of antibodies in virus-exposed individuals. Here we review the latest knowledge on humoral immune responses to SARS-CoV-2 infection, along with the benefits and limitations of currently available commercial and laboratory-based serological assays. We also highlight important serological considerations, such as antibody expression levels, stability and neutralization dynamics, as well as cross-reactivity and possible immunological back-boosting by seasonal coronaviruses. The ability to accurately detect, measure and characterize the various antibodies specific to SARS-CoV-2 is necessary for vaccine development, manage risk and exposure for healthcare and at-risk workers, and for monitoring reinfections with genetic variants and new strains of the virus. Having a thorough understanding of the benefits and cautions of standardized serological testing at a community level remains critically important in the design and implementation of future vaccination campaigns, epidemiological models of immunity, and public health measures that rely heavily on up-to-date knowledge of transmission dynamics.

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