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Proteomic screening of antigenic proteins from the hard tick, Haemaphysalis longicornis (Acari: Ixodidae).

Kim YH, Slam MS, You MJ - Korean J. Parasitol. (2015)

Bottom Line: The proteome of the partially fed female had a larger number of lower molecular weight proteins than that of the unfed female tick.These findings indicate that most of the whole body components of these ticks are non-immunogenic.The data reported here will provide guidance in the identification of antigenic proteins to prevent infestation and diseases transmitted by H. longicornis.

View Article: PubMed Central - PubMed

Affiliation: Department of Veterinary Parasitology , College of Veterinary Medicine and Biosafety Research Centre, Chonbuk National University, Jeonju 561-756, Korea.

ABSTRACT
Proteomic tools allow large-scale, high-throughput analyses for the detection, identification, and functional investigation of proteome. For detection of antigens from Haemaphysalis longicornis, 1-dimensional electrophoresis (1-DE) quantitative immunoblotting technique combined with 2-dimensional electrophoresis (2-DE) immunoblotting was used for whole body proteins from unfed and partially fed female ticks. Reactivity bands and 2-DE immunoblotting were performed following 2-DE electrophoresis to identify protein spots. The proteome of the partially fed female had a larger number of lower molecular weight proteins than that of the unfed female tick. The total number of detected spots was 818 for unfed and 670 for partially fed female ticks. The 2-DE immunoblotting identified 10 antigenic spots from unfed females and 8 antigenic spots from partially fed females. Matrix Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry (MALDI-TOF) of relevant spots identified calreticulin, putative secreted WC salivary protein, and a conserved hypothetical protein from the National Center for Biotechnology Information and Swiss Prot protein sequence databases. These findings indicate that most of the whole body components of these ticks are non-immunogenic. The data reported here will provide guidance in the identification of antigenic proteins to prevent infestation and diseases transmitted by H. longicornis.

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Representative 2-DE images of the whole body proteins from unfed female ticks of H. longicornis. (A) Silver stained gel. (B) Immunoblot probed with polyclonal antibodies against H. longicornis. Proteins (240 μg) were focused on pH 4-7 IPG strips (13 cm) and were then separated by SDS/12.5% PAGE. Reference molecular masses are indicated on the right. 2-DE immunoblot shows the antigenic spots revealed by a pool of sera from rabbits experimentally infested with H. longicornis. The proteins of interest are marked in the image (rectangles). (C, D) Magnifications of panels A and B. In D, antigenic spots are indicated by black numbered circles and are equivalent to the proteins identified in panel C.
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f4-kjp-53-1-85: Representative 2-DE images of the whole body proteins from unfed female ticks of H. longicornis. (A) Silver stained gel. (B) Immunoblot probed with polyclonal antibodies against H. longicornis. Proteins (240 μg) were focused on pH 4-7 IPG strips (13 cm) and were then separated by SDS/12.5% PAGE. Reference molecular masses are indicated on the right. 2-DE immunoblot shows the antigenic spots revealed by a pool of sera from rabbits experimentally infested with H. longicornis. The proteins of interest are marked in the image (rectangles). (C, D) Magnifications of panels A and B. In D, antigenic spots are indicated by black numbered circles and are equivalent to the proteins identified in panel C.

Mentions: As shown in Fig. 4B and D, the 2-DE immunoblot analysis of the proteins from unfed female ticks with sera from rabbits infested by H. longicornis revealed around 10 major spots in the range of pI 4.0-6.5 and 35-100 kDa. Matching of the immunoblot with its homologous silver-stained gel allowed us to localize these 10 major antigenic spots in the 2-DE gels (Fig. 4C, D). The 2-DE immunoblot analysis of the proteins from partially fed female ticks with sera from rabbits infested by H. longicornis revealed around 8 major spots in a narrow range of pI (4.0-5.0) and molecular weight (16-35 kDa), and some minor and poorly resolved spots in the range of pI 5.5-6.0 and 25-30 kDa (Fig. 5B, D). Matching of the immunoblot with its homologous silver-stained gel allowed us to localize the 8 major antigenic spots in the 2-DE gels (Fig. 5C, D).


Proteomic screening of antigenic proteins from the hard tick, Haemaphysalis longicornis (Acari: Ixodidae).

Kim YH, Slam MS, You MJ - Korean J. Parasitol. (2015)

Representative 2-DE images of the whole body proteins from unfed female ticks of H. longicornis. (A) Silver stained gel. (B) Immunoblot probed with polyclonal antibodies against H. longicornis. Proteins (240 μg) were focused on pH 4-7 IPG strips (13 cm) and were then separated by SDS/12.5% PAGE. Reference molecular masses are indicated on the right. 2-DE immunoblot shows the antigenic spots revealed by a pool of sera from rabbits experimentally infested with H. longicornis. The proteins of interest are marked in the image (rectangles). (C, D) Magnifications of panels A and B. In D, antigenic spots are indicated by black numbered circles and are equivalent to the proteins identified in panel C.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4384799&req=5

f4-kjp-53-1-85: Representative 2-DE images of the whole body proteins from unfed female ticks of H. longicornis. (A) Silver stained gel. (B) Immunoblot probed with polyclonal antibodies against H. longicornis. Proteins (240 μg) were focused on pH 4-7 IPG strips (13 cm) and were then separated by SDS/12.5% PAGE. Reference molecular masses are indicated on the right. 2-DE immunoblot shows the antigenic spots revealed by a pool of sera from rabbits experimentally infested with H. longicornis. The proteins of interest are marked in the image (rectangles). (C, D) Magnifications of panels A and B. In D, antigenic spots are indicated by black numbered circles and are equivalent to the proteins identified in panel C.
Mentions: As shown in Fig. 4B and D, the 2-DE immunoblot analysis of the proteins from unfed female ticks with sera from rabbits infested by H. longicornis revealed around 10 major spots in the range of pI 4.0-6.5 and 35-100 kDa. Matching of the immunoblot with its homologous silver-stained gel allowed us to localize these 10 major antigenic spots in the 2-DE gels (Fig. 4C, D). The 2-DE immunoblot analysis of the proteins from partially fed female ticks with sera from rabbits infested by H. longicornis revealed around 8 major spots in a narrow range of pI (4.0-5.0) and molecular weight (16-35 kDa), and some minor and poorly resolved spots in the range of pI 5.5-6.0 and 25-30 kDa (Fig. 5B, D). Matching of the immunoblot with its homologous silver-stained gel allowed us to localize the 8 major antigenic spots in the 2-DE gels (Fig. 5C, D).

Bottom Line: The proteome of the partially fed female had a larger number of lower molecular weight proteins than that of the unfed female tick.These findings indicate that most of the whole body components of these ticks are non-immunogenic.The data reported here will provide guidance in the identification of antigenic proteins to prevent infestation and diseases transmitted by H. longicornis.

View Article: PubMed Central - PubMed

Affiliation: Department of Veterinary Parasitology , College of Veterinary Medicine and Biosafety Research Centre, Chonbuk National University, Jeonju 561-756, Korea.

ABSTRACT
Proteomic tools allow large-scale, high-throughput analyses for the detection, identification, and functional investigation of proteome. For detection of antigens from Haemaphysalis longicornis, 1-dimensional electrophoresis (1-DE) quantitative immunoblotting technique combined with 2-dimensional electrophoresis (2-DE) immunoblotting was used for whole body proteins from unfed and partially fed female ticks. Reactivity bands and 2-DE immunoblotting were performed following 2-DE electrophoresis to identify protein spots. The proteome of the partially fed female had a larger number of lower molecular weight proteins than that of the unfed female tick. The total number of detected spots was 818 for unfed and 670 for partially fed female ticks. The 2-DE immunoblotting identified 10 antigenic spots from unfed females and 8 antigenic spots from partially fed females. Matrix Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry (MALDI-TOF) of relevant spots identified calreticulin, putative secreted WC salivary protein, and a conserved hypothetical protein from the National Center for Biotechnology Information and Swiss Prot protein sequence databases. These findings indicate that most of the whole body components of these ticks are non-immunogenic. The data reported here will provide guidance in the identification of antigenic proteins to prevent infestation and diseases transmitted by H. longicornis.

Show MeSH
Related in: MedlinePlus