Creative Biolabs Released New Antibody Sequencing Technique

Antibody sequencing is a technique that determines the amino acid sequence of an antibody.
Discovering the structures and functions of antibodies in living organism is an important tool
in understanding cellular processes, which allow drugs that target specific metabolic
pathways to be invented more easily.

This next generation antibody sequencing technique has now become a routine measurement
that can be completed within 10 working days for IgGs. The method is applicable for all
antibody formats, namely IgMs, fluorochrome conjugates, immobilized antibodies and
mixtures. Creative Biolabs, an emerging qualified service provider in this field, provides
researchers their unique technique in antibody sequencing.

With the development of biotechnology, scientists of Creative Biolabs have developed a

novel strategy to determine the complete sequence of an antibody with unparalleled speed and
accuracy. The technique which combines all the advantages of existing approaches is named
"Database Assisted Shotgun Sequencing" (DASS).

In the first step, the antibody is fragmented to peptides by a special technique, which
generates up to 5000 different peptides per chain. This set of peptides will be analyzed by
high end mass spectrometers to generate extensive sequence information. MS/MS spectra are
then de novo sequenced by the latest algorithms and matched against a database with related
sequences. Special "in house" data mining tools allow Creative Biolabs to extract the
sequence information from ten thousands of MS/MS spectra within hours.

De Novo Sequencing of the CDR3 Region

While the CDR3 of the light chain is mostly encoded by the germline sequences, the CDR3 of
the heavy chain is usually not available in databases. It is encoded by the so called
D-segments, but these are modified by nucleases and terminal transferases. Typically, only
1-4 AA of a D-segment remain in the matured antibody. The rest of the D-segment is
"artificial" and has to go through phage display techniques. Their method generates many
overlapping peptides during the fragmentation process, enabling them to sequence very long
stretches of unknown amino acids. The high quality of MS/MS spectra in combination with
intelligent data mining, allows them to read the CDR3 like a book. The technique is so
powerful, that they were able to sequence a 20 kDa protein, which had no homologue in the
database.

Sequencing of the V and J and C segments by "Database Assisted Shotgun Sequencing"

The V and J and C gene segments of antibodies are available in public databases. During the
maturation of an antibody, the B-cell introduces hypermutations into the sequence to optimize
the affinity. Creative Biolabs's mapping algorithm is error tolerant and can match the
"mutated" peptides to the corresponding germline reliably.
Because of the great number of peptides, Creative Biolabs is able to get sequence information
for ever bond in the antibody. Typically, 20-70 different MS/MS spectra are generated for
each amino acid (AA) position. Hence, even the hardest sequences of proline and arginine
rich peptides can be resolved.

About Creative Biolabs

Creative Biolabs is specialized in providing custom biotechnology and pharmaceutical

services that cover the full scope of biotechnology needs of early drug discovery and
development, including antibody production , membrane protein preparation, phage display
techniques , etc.

Source:
http://www.bio-itworld.com/Press-Release/Creative-Biolabs-Released-New-Antibody
-Sequencing-Technique/